Diaryl substituted 5,5-fused ring compounds as C5aR inhibitors

ABSTRACT

The present disclosure provides, inter alia, Compounds of Formula (I) 
                         
or pharmaceutically acceptable salts thereof that are modulators of the C5a receptor. Also provided are pharmaceutical compositions and methods of use including the treatment of diseases or disorders involving pathologic activation from C5a and non-pharmaceutical applications.

CROSS-REFERENCES TO RELATED APPLICATIONS

This application is an application claiming benefit under 35 U.S.C. §119(e) of U.S. Provisional Application No. 62/609,844 filed Dec. 22,2017, which is herein incorporated by reference in its entirety.

STATEMENT AS TO RIGHTS TO INVENTIONS MADE UNDER FEDERALLY SPONSOREDRESEARCH AND DEVELOPMENT

NOT APPLICABLE

REFERENCE TO A “SEQUENCE LISTING,” A TABLE, OR A COMPUTER PROGRAMLISTING APPENDIX SUBMITTED ON A COMPACT DISK

NOT APPLICABLE

BACKGROUND OF THE INVENTION

The complement system plays a central role in the clearance of immunecomplexes and in immune responses to infectious agents, foreignantigens, virus infected cells and tumor cells. Inappropriate orexcessive activation of the complement system can lead to harmful, andeven potentially life-threatening consequences due to severeinflammation and resulting tissue destruction. These consequences areclinically manifested in various disorders including septic shock;myocardial, as well as, intestinal ischemia/reperfusion injury; graftrejection; organ failure; nephritis; pathological inflammation; andautoimmune diseases.

The complement system is composed of a group of proteins that arenormally present in the serum in an inactive state. Activation of thecomplement system encompasses mainly three distinct pathways, i.e., theclassical, the alternative, and the lectin pathway (V. M. Holers, InClinical Immunology: Principles and Practice, ed. R. R. Rich, MosbyPress; 1996, 363-391): 1) The classical pathway is acalcium/magnesium-dependent cascade, which is normally activated by theformation of antigen-antibody complexes. It can also be activated in anantibody-independent manner by the binding of C-reactive protein,complexed with ligand, and by many pathogens including gram-negativebacteria. 2) The alternative pathway is a magnesium-dependent cascadewhich is activated by deposition and activation of C3 on certainsusceptible surfaces (e.g. cell wall polysaccharides of yeast andbacteria, and certain biopolymer materials). 3) The lectin pathwayinvolves the initial binding of mannose-binding lectin and thesubsequent activation of C2 and C4, which are common to the classicalpathway (Matsushita, M. et al., J. Exp. Med. 176: 1497-1502 (1992);Suankratay, C. et al., J. Immunol. 160: 3006-3013 (1998)).

The activation of the complement pathway generates biologically activefragments of complement proteins, e.g. C3a, C4a and C5a anaphylatoxinsand C5b-9 membrane attack complexes (MAC), all which mediateinflammatory responses by affecting leukocyte chemotaxis; activatingmacrophages, neutrophils, platelets, mast cells and endothelial cells;and increasing vascular permeability, cytolysis and tissue injury.

Complement C5a is one of the most potent proinflammatory mediators ofthe complement system. (The anaphylactic C5a peptide is 100 times morepotent, on a molar basis, in eliciting inflammatory responses than C3a.)C5a is the activated form of C5 (190 kD, molecular weight). C5a ispresent in human serum at approximately 80 μg/ml (Kohler, P. F. et al.,J. Immunol. 99: 1211-1216 (1967)). It is composed of two polypeptidechains, α and β, with approximate molecular weights of 115 kD and 75 kD,respectively (Tack, B. F. et al., Biochemistry 18: 1490-1497 (1979)).Biosynthesized as a single-chain promolecule, C5 is enzymaticallycleaved into a two-chain structure during processing and secretion.After cleavage, the two chains are held together by at least onedisulphide bond as well as noncovalent interactions (Ooi, Y. M. et al.,J. Immunol. 124: 2494-2498(1980)).

C5 is cleaved into the C5a and C5b fragments during activation of thecomplement pathways. The convertase enzymes responsible for C5activation are multi-subunit complexes of C4b, C2a, and C3b for theclassical pathway and of (C3b)₂, Bb, and P for the alternative pathway(Goldlust, M. B. et al., J. Immunol. 113: 998-1007 (1974); Schreiber, R.D. et al, Proc. Natl. Acad. Sci. 75: 3948-3952 (1978)). C5 is activatedby cleavage at position 74-75 (Arg-Leu) in the α-chain. Afteractivation, the 11.2 kD, 74 amino acid peptide C5a from theamino-terminus portion of the α-chain is released. Both C5a and C3a arepotent stimulators of neutrophils and monocytes (Schindler, R. et al.,Blood 76: 1631-1638 (1990); Haeffner-Cavaillon, N. et al., J. Immunol.138: 794-700 (1987); Cavaillon, J. M. et al., Eur. J. Immunol. 20:253-257 (1990)).

In addition to its anaphylatoxic properties, C5a induces chemotacticmigration of neutrophils (Ward, P. A. et al., J. Immunol. 102: 93-99(1969)), eosinophils (Kay, A. B. et al., Immunol. 24: 969-976 (1973)),basophils (Lett-Brown, M. A. et al., J. Immunol. 117: 246-252 1976)),and monocytes (Snyderman, R. et al., Proc. Soc. Exp. Biol. Med. 138:387-390 1971)). Both C5a and C5b-9 activate endothelial cells to expressadhesion molecules essential for sequestration of activated leukocytes,which mediate tissue inflammation and injury (Foreman, K. E. et al., J.Clin. Invest. 94: 1147-1155 (1994); Foreman, K. E. et al., Inflammation20: 1-9 (1996); Rollins, S. A. et al., Transplantation 69: 1959-1967(2000)). C5a also mediates inflammatory reactions by causing smoothmuscle contraction, increasing vascular permeability, inducing basophiland mast cell degranulation and inducing release of lysosomal proteasesand oxidative free radicals (Gerard, C. et al., Ann. Rev. Immunol. 12:775-808 (1994)). Furthermore, C5a modulates the hepatic acute-phase geneexpression and augments the overall immune response by increasing theproduction of TNF-α, IL-1-β, IL-6, IL-8, prostaglandins and leukotrienes(Lambris, J. D. et al., In: The Human Complement System in Health andDisease, Volanakis, J. E. ed., Marcel Dekker, New York, pp. 83-118).

The anaphylactic and chemotactic effects of C5a are believed to bemediated through its interaction with the C5a receptor. The human C5areceptor (C5aR) is a 52 kD membrane bound G protein-coupled receptor,and is expressed on neutrophils, monocytes, basophils, eosinophils,hepatocytes, lung smooth muscle and endothelial cells, and renalglomerular tissues (Van-Epps, D. E. et al., J. Immunol. 132: 2862-2867(1984); Haviland, D. L. et al., J. Immunol. 154:1861-1869 (1995);Wetsel, R. A., Immunol. Leff. 44: 183-187 (1995); Buchner, R. R. et al.,J. Immunol. 155: 308-315 (1995); Chenoweth, D. E. et al., Proc. Natl.Acad. Sci. 75: 3943-3947 (1978); Zwirner, J. et al., Mol. Immunol.36:877-884 (1999)). The ligand-binding site of C5aR is complex andconsists of at least two physically separable binding domains. One bindsthe C5a amino terminus (amino acids 1-20) and disulfide-linked core(amino acids 21-61), while the second binds the C5a carboxy-terminal end(amino acids 62-74) (Wetsel, R. A., Curr. Opin. Immunol. 7: 48-53(1995)).

C5a plays important roles in inflammation and tissue injury. Incardiopulmonary bypass and hemodialysis, C5a is formed as a result ofactivation of the alternative complement pathway when human blood makescontact with the artificial surface of the heart-lung machine or kidneydialysis machine (Howard, R. J. et al., Arch. Surg. 123: 1496-1501(1988); Kirklin, J. K. et al., J. Cardiovasc. Surg. 86: 845-857 (1983);Craddock, P. R. et al., N. Engl. J. Med. 296: 769-774 (1977)). C5acauses increased capillary permeability and edema, bronchoconstriction,pulmonary vasoconstriction, leukocyte and platelet activation andinfiltration to tissues, in particular the lung (Czermak, B. J. et al.,J. Leukoc. Biol. 64: 40-48 (1998)). Administration of an anti-05amonoclonal antibody was shown to reduce cardiopulmonary bypass andcardioplegia-induced coronary endothelial dysfunction (Tofukuji, M. etal., J. Thorac. Cardiovasc. Surg. 116: 1060-1068 (1998)).

C5a is also involved in acute respiratory distress syndrome (ARDS),Chronic Obstructive Pulmonary Disorder (COPD) and multiple organ failure(MOF) (Hack, C. E. et al., Am. J. Med. 1989: 86: 20-26; Hammerschmidt DE et al. Lancet 1980; 1: 947-949; Heideman M. et al. J. Trauma 1984; 4:1038-1043; Marc, M M, et al., Am. J. Respir. Cell and Mol. Biol., 2004:31: 216-219). C5a augments monocyte production of two importantpro-inflammatory cytokines, TNF-α and IL-1. C5a has also been shown toplay an important role in the development of tissue injury, andparticularly pulmonary injury, in animal models of septic shock(Smedegard G et al. Am. J. Pathol. 1989; 135: 489-497; Markus, S., etal., FASEB Journal (2001), 15: 568-570). In sepsis models using rats,pigs and non-human primates, anti-05a antibodies administered to theanimals before treatment with endotoxin or E. coli resulted in decreasedtissue injury, as well as decreased production of IL-6 (Smedegard, G. etal., Am. J. Pathol. 135: 489-497 (1989); Hopken, U. et al., Eur. J.Immunol. 26: 1103-1109 (1996); Stevens, J. H. et al., J. Clin. Invest.77: 1812-1816 (1986)). More importantly, blockade or C5a with anti-05apolyclonal antibodies has been shown to significantly improve survivalrates in a caecal ligation/puncture model of sepsis in rats (Czermak, B.J. et al., Nat. Med. 5: 788-792 (1999)). This model share many aspectsof the clinical manifestation of sepsis in humans. (Parker, S. J. etal., Br. J. Surg. 88: 22-30 (2001)). In the same sepsis model, anti-05aantibodies were shown to inhibit apoptosis of thymocytes (Guo, R. F. etal., J. Clin. Invest. 106: 1271-1280 (2000)) and prevent MOF(Huber-Lang, M. et al., J. Immunol. 166: 1193-1199 (2001)). Anti-C5aantibodies were also protective in a cobra venom factor model of lunginjury in rats, and in immune complex-induced lung injury (Mulligan, M.S. et al. J. Clin. Invest. 98: 503-512 (1996)). The importance of C5a inimmune complex-mediated lung injury was later confirmed in mice (Bozic,C. R. et al., Science 26: 1103-1109 (1996)).

C5a is found to be a major mediator in myocardial ischemia-reperfusioninjury. Complement depletion reduced myocardial infarct size in mice(Weisman, H. F. et al., Science 249: 146-151 (1990)), and treatment withanti-05a antibodies reduced injury in a rat model of hindlimbischemia-reperfusion (Bless, N. M. et al., Am. J. Physiol. 276: L57-L63(1999)). Reperfusion injury during myocardial infarction was alsomarkedly reduced in pigs that were retreated with a monoclonal anti-05aIgG (Amsterdam, E. A. et al., Am. J. Physiol. 268:H448-H457 (1995)). Arecombinant human C5aR antagonist reduces infarct size in a porcinemodel of surgical revascularization (Riley, R. D. et al., J. Thorac.Cardiovasc. Surg. 120: 350-358 (2000)).

C5a driven neutrophils also contribute to many bullous diseases (e.g.,bullous pemphigoid, pemphigus vulgaris and pemphigus foliaceus). Theseare chronic and recurring inflammatory disorders clinicallycharacterized by sterile blisters that appear in the sub-epidermal spaceof the skin and mucosa. While autoantibodies to keratinocytes located atthe cutaneous basement membranes are believed to underlie the detachmentof epidermal basal keratinocytes from the underlying basement membrane,blisters are also characterized by accumulation of neutrophils in boththe upper dermal layers and within the blister cavities. In experimentalmodels a reduction of neutrophils or absence of complement (total orC5-selective) can inhibit formation of sub-epidermal blisters, even inthe presence of high auto-antibody titers.

Complement levels are elevated in patients with rheumatoid arthritis(Jose, P. J. et al., Ann. Rheum. Dis. 49: 747-752 (1990); Grant, E. P.,et al., J. of Exp. Med., 196(11): 1461-1471, (2002)), lupus nephritis(Bao, L., et al., Eur. J. of Immunol., 35(8), 2496-2506, (2005)) andsystemic lupus erythematosus (SLE) (Porcel, J. M. et al., Clin. Immunol.Immunopathol. 74: 283-288 (1995)). C5a levels correlate with theseverity of the disease state. Collagen-induced arthritis in mice andrats resembles the rheumatoid arthritic disease in human. Mice deficientin the C5a receptor demonstrated a complete protection from arthritisinduced by injection of monoclonal anti-collagen Abs (Banda, N. K., etal., J. of Immunol., 2003, 171: 2109-2115). Therefore, inhibition of C5aand/or C5a receptor (C5aR) could be useful in treating these chronicdiseases.

The complement system is believed to be activated in patients withinflammatory bowel disease (IBD) and is thought to play a role in thedisease pathogenesis. Activated complement products were found at theluminal face of surface epithelial cells, as well as in the muscularismucosa and submucosal blood vessels in IBD patients (Woodruff, T. M., etal., J of Immunol., 2003, 171: 5514-5520).

C5aR expression is upregulated on reactive astrocytes, microglia, andendothelial cells in an inflamed human central nervous system (Gasque,P. et al., Am. J. Pathol. 150: 31-41 (1997)). C5a might be involved inneurodegenerative diseases, such as Alzheimer disease (Mukherjee, P. etal., J. Neuroimmunol. 105: 124-130 (2000); O'Barr, S. et al., J.Neuroimmunol. (2000) 105: 87-94; Farkas, I., et al. J. Immunol. (2003)170:5764-5771), Parkinson's disease, Pick disease and transmissiblespongiform encephalopathies. Activation of neuronal C5aR may induceapoptosis (Farkas I et al. J. Physiol. 1998; 507: 679-687). Therefore,inhibition of C5a and/or C5aR could also be useful in treatingneurodegenerative diseases.

There is some evidence that C5a production worsens inflammationassociated with atopic dermatitis (Neuber, K., et al., Immunology73:83-87, (1991)), and chronic urticaria (Kaplan, A. P., J. AllergyClin. Immunol. 114; 465-474, (2004).

Psoriasis is now known to be a T cell-mediated disease (Gottlieb, E. L.et al., Nat. Med. 1: 442-447 (1995)). However, neutrophils and mastcells may also be involved in the pathogenesis of the disease (Terui, T.et al., Exp. Dermatol. 9: 1-10; 2000); Werfel, T. et al., Arch.Dermatol. Res. 289: 83-86 (1997)). Neutrophil accumulation under thestratum corneum is observed in the highly inflamed areas of psoriaticplaques, and psoriatic lesion (scale) extracts contain highly elevatedlevels of C5a and exhibit potent chemotactic activity towardsneutrophils, an effect that can be inhibited by addition of a C5aantibody. T cells and neutrophils are chemo-attracted by C5a (Nataf, S.et al., J. Immunol. 162: 4018-4023 (1999); Tsuji, R. F. et al., J.Immunol. 165: 1588-1598 (2000); Cavaillon, J. M. et al., Eur. J.Immunol. 20: 253-257 (1990)). Additionally expression of C5aR has beendemonstrated in plasmacytoid dendritic cells (pDC) isolated from lesionsof cutaneous lupus erythematous and these cells were shown to displaychemotactic behavior towards C5a, suggesting that blockade of C5aR onpDC might be efficacious in reducing pDC infiltration into inflamed skinin both SLE and psoriasis. Therefore C5a could be an importanttherapeutic target for treatment of psoriasis.

Immunoglobulin G-containing immune complexes (IC) contribute to thepathophysiology in a number of autoimmune diseases, such as systemiclupus erthyematosus, rheumatoid arthritis, Sjogren's disease,Goodpasture's syndrome, and hypersensitivity pneumonitis (Madaio, M. P.,Semin. Nephrol. 19: 48-56 (1999); Korganow, A. S. et al., Immunity 10:451-459 (1999); Bolten, W. K., Kidney Int. 50: 1754-1760 (1996); Ando,M. et al., Curr. Opin. Pulm. Med. 3: 391-399 (1997)). These diseases arehighly heterogeneous and generally affect one or more of the followingorgans: skin, blood vessels, joints, kidneys, heart, lungs, nervoussystem and liver (including cirrhosis and liver fibrosis). The classicalanimal model for the inflammatory response in these IC diseases is theArthus reaction, which features the infiltration of polymorphonuclearcells, hemorrhage, and plasma exudation (Arthus, M., C.R. Soc. Biol. 55:817-824 (1903)). Recent studies show that C5aR deficient mice areprotected from tissue injury induced by IC (Kohl, J. et al., Mol.Immunol. 36: 893-903 (1999); Baumann, U. et al., J. Immunol. 164:1065-1070 (2000)). The results are consistent with the observation thata small peptidic anti-05aR antagonist inhibits the inflammatory responsecaused by IC deposition (Strachan, A. J. et al., J. Immunol. 164:6560-6565 (2000)). Together with its receptor, C5a plays an importantrole in the pathogenesis of IC diseases. Inhibitors of C5a and C5aRcould be useful to treat these diseases.

DESCRIPTION OF RELATED ART

Non-peptide based C5a receptor antagonist have been reported as beingeffective for treating endotoxic shock in rats (Stracham, A. J., et al.,J. of Immunol. (2000), 164(12): 6560-6565); and for treating IBD in arat model (Woodruff, T. M., et al., J. of Immunol., 2003, 171:5514-5520). Non-peptide based C5a receptor modulators also have beendescribed in the patent literature by Neurogen Corporation, (e.g.,WO2004/043925, WO2004/018460, WO2005/007087, WO03/082826, WO03/08828,WO02/49993, WO03/084524); Dompe S. P. A. (WO02/029187); The Universityof Queenland (WO2004/100975); and ChemoCentryx (WO2010/075257).

There is considerable experimental evidence in the literature thatimplicates increased levels of C5a with a number of diseases anddisorders, in particular in autoimmune and inflammatory diseases anddisorders. Thus, there remains a need in the art for new small organicmolecule modulators, e.g., agonists, preferably antagonists, partialagonists, of the C5a receptor (C5aR) that are useful for inhibitingpathogenic events, e.g., chemotaxis, associated with increased levelsanaphylatoxin activity. The present invention fulfills this and otherneeds.

BRIEF SUMMARY OF THE INVENTION

In one aspect, the present invention provide compounds of Formula (I):

or a pharmaceutically acceptable salt thereof, wherein the symbols,letters and subscripts n, m, a, b, e, X¹, R¹, R^(2a), R^(2b), R³, R⁴ andR⁵ have the meanings provided in the description below.

In addition to the compounds provided herein, the present inventionfurther provides pharmaceutical compositions containing one or more ofthese compounds, as well as methods for the use of these compounds intherapeutic methods, primarily to treat diseases associated C5asignaling activity.

In yet another aspect, the present invention provides methods ofdiagnosing disease in an individual. In these methods, the compoundsprovided herein are administered in labeled form to a subject, followedby diagnostic imaging to determine the presence or absence of C5aRand/or the localization of cells expressing a C5aR receptor. In arelated aspect, a method of diagnosing disease is carried out bycontacting a tissue or blood sample with a labeled compound as providedherein and determining the presence, absence, amount, or localization ofC5aR in the sample.

BRIEF DESCRIPTION OF THE DRAWINGS

NOT APPLICABLE

DETAILED DESCRIPTION OF THE INVENTION I. Abbreviation and Definitions

The term “alkyl”, by itself or as part of another substituent, means,unless otherwise stated, a straight or branched chain hydrocarbonradical, having the number of carbon atoms designated (i.e. C₁₋₈ meansone to eight carbons). Examples of alkyl groups include methyl, ethyl,n-propyl, isopropyl, n-butyl, t-butyl, isobutyl, sec-butyl, n-pentyl,n-hexyl, n-heptyl, n-octyl, and the like. The term “alkenyl” refers toan unsaturated alkyl group having one or more double bonds. Similarly,the term “alkynyl” refers to an unsaturated alkyl group having one ormore triple bonds. Examples of such unsaturated alkyl groups includevinyl, 2-propenyl, crotyl, 2-isopentenyl, 2-(butadienyl), isobutenyl,2,4-pentadienyl, 3-(1,4-pentadienyl), ethynyl, 1- and 3-propynyl,3-butynyl, and the higher homologs and isomers. The term “cycloalkyl”refers to hydrocarbon rings having the indicated number of ring atoms(e.g., C₃₋₆cycloalkyl) and being fully saturated or having no more thanone double bond between ring vertices. “Cycloalkyl” is also meant torefer to bicyclic and polycyclic hydrocarbon rings such as, for example,bicyclo[2.2.1]heptane, bicyclo[2.2.2]octane, etc. The term“heterocycloalkyl” refers to a cycloalkyl group that contain from one tofive heteroatoms selected from N, O, and S, wherein the nitrogen andsulfur atoms are optionally oxidized, and the nitrogen atom(s) areoptionally quaternized. The heterocycloalkyl may be a monocyclic, abicyclic or a polycylic ring system. Non limiting examples ofheterocycloalkyl groups include pyrrolidine, imidazolidine,pyrazolidine, butyrolactam, valerolactam, imidazolidinone, hydantoin,dioxolane, phthalimide, piperidine, 1,4-dioxane, morpholine,thiomorpholine, thiomorpholine-S-oxide, thiomorpholine-S,S-oxide,piperazine, pyran, pyridone, 3-pyrroline, thiopyran, pyrone,tetrahydrofuran, tetrhydrothiophene, quinuclidine, and the like. Aheterocycloalkyl group can be attached to the remainder of the moleculethrough a ring carbon or a heteroatom.

The term “alkylene” by itself or as part of another substituent means adivalent radical derived from an alkane, as exemplified by—CH₂CH₂CH₂CH₂—. Typically, an alkyl (or alkylene) group will have from 1to 24 carbon atoms, with those groups having 10 or fewer carbon atomsbeing preferred in the present invention. A “lower alkyl” or “loweralkylene” is a shorter chain alkyl or alkylene group, generally havingfour or fewer carbon atoms. Similarly, “alkenylene” and “alkynylene”refer to the unsaturated forms of “alkylene” having double or triplebonds, respectively.

The term “heteroalkyl,” by itself or in combination with another term,means, unless otherwise stated, a stable straight or branched chain, orcyclic hydrocarbon radical, or combinations thereof, consisting of thestated number of carbon atoms and from one to three heteroatoms selectedfrom the group consisting of O, N, Si and S, and wherein the nitrogenand sulfur atoms may optionally be oxidized and the nitrogen heteroatommay optionally be quaternized. The heteroatom(s) O, N and S may beplaced at any interior position of the heteroalkyl group. The heteroatomSi may be placed at any position of the heteroalkyl group, including theposition at which the alkyl group is attached to the remainder of themolecule. Examples include —CH₂—CH₂—O—CH₃, —CH₂—CH₂—NH—CH₃,—CH₂—CH₂—N(CH₃)—CH₃, —CH₂—S—CH₂—CH₃, —CH₂—CH₂, —S(O)—CH₃,—CH₂—CH₂—S(O)₂—CH₃, —CH═CH—O—CH₃, —Si(CH₃)₃, —CH₂—CH═N—OCH₃, and—CH═CH—N(CH₃)—CH₃. Up to two heteroatoms may be consecutive, such as,for example, —CH₂—NH—OCH₃ and —CH₂—O—Si(CH₃)₃. Similarly, the terms“heteroalkenyl” and “heteroalkynyl” by itself or in combination withanother term, means, unless otherwise stated, an alkenyl group oralkynyl group, respectively, that contains the stated number of carbonsand having from one to three heteroatoms selected from the groupconsisting of O, N, Si and S, and wherein the nitrogen and sulfur atomsmay optionally be oxidized and the nitrogen heteroatom may optionally bequaternized. The heteroatom(s) O, N and S may be placed at any interiorposition of the heteroalkyl group.

The term “heteroalkylene” by itself or as part of another substituentmeans a divalent radical, saturated or unsaturated or polyunsaturated,derived from heteroalkyl, as exemplified by —CH₂—CH₂—S—CH₂CH₂— and—CH₂—S—CH₂—CH₂—NH—CH₂—, —O—CH₂—CH═CH—, —CH₂—CH═C(H)CH₂—O—CH₂— and—S—CH₂—C—. For heteroalkylene groups, heteroatoms can also occupy eitheror both of the chain termini (e.g., alkyleneoxy, alkylenedioxy,alkyleneamino, alkylenediamino, and the like).

The terms “alkoxy,” “alkylamino” and “alkylthio” (or thioalkoxy) areused in their conventional sense, and refer to those alkyl groupsattached to the remainder of the molecule via an oxygen atom, an aminogroup, or a sulfur atom, respectively. Additionally, for dialkylaminogroups, the alkyl portions can be the same or different and can also becombined to form a 3-7 membered ring with the nitrogen atom to whicheach is attached. Accordingly, a group represented as —NR^(a)R^(b) ismeant to include piperidinyl, pyrrolidinyl, morpholinyl, azetidinyl andthe like.

The term “hydroxyalkyl” is used in its conventional sense, and refers tobranched or straight chain alkyl group substituted with at least onehydroxyl group. The hydroxyl group may be at any position in the alkylgroup. For example, the term “C₁₋₄ hydroxylalkyl” is meant to includehydroxymethyl, hydroxyethyl, hydroxypropyl, hydroxyisopropyl, and thelike.

The terms “halo” or “halogen,” by themselves or as part of anothersubstituent, mean, unless otherwise stated, a fluorine, chlorine,bromine, or iodine atom. Additionally, terms such as “haloalkyl,” aremeant to include monohaloalkyl and polyhaloalkyl. For example, the term“C₁₋₄ haloalkyl” is mean to include trifluoromethyl,2,2,2-trifluoroethyl, 4-chlorobutyl, 3-bromopropyl, and the like.

The term “aryl” means, unless otherwise stated, a polyunsaturated,typically aromatic, hydrocarbon group which can be a single ring ormultiple rings (up to three rings) which are fused together or linkedcovalently. The term “heteroaryl” refers to aryl groups (or rings) thatcontain from one to five heteroatoms selected from N, O, and S, whereinthe nitrogen and sulfur atoms are optionally oxidized, and the nitrogenatom(s) are optionally quaternized. A heteroaryl group can be attachedto the remainder of the molecule through a heteroatom. Non-limitingexamples of aryl groups include phenyl, naphthyl and biphenyl, whilenon-limiting examples of heteroaryl groups include pyridyl, pyridazinyl,pyrazinyl, pyrimindinyl, triazinyl, quinolinyl, quinoxalinyl,quinazolinyl, cinnolinyl, phthalaziniyl, benzotriazinyl, purinyl,benzimidazolyl, benzopyrazolyl, benzooxazolyl, benzotriazolyl,benzisoxazolyl, isobenzofuryl, isoindolyl, indolizinyl, benzotriazinyl,thienopyridinyl, thienopyrimidinyl, pyrazolopyrimidinyl, pyrrolopyridyl,imidazopyridines, benzothiaxolyl, benzofuranyl, benzothienyl, indolyl,quinolyl, isoquinolyl, isothiazolyl, pyrazolyl, indazolyl, pteridinyl,imidazolyl, triazolyl, tetrazolyl, oxazolyl, isoxazolyl, thiadiazolyl,pyrrolyl, thiazolyl, furyl, thienyl and the like. Substituents for eachof the above noted aryl and heteroaryl ring systems are selected fromthe group of acceptable substituents described below.

The term “pharmaceutically acceptable salts” is meant to include saltsof the active compounds which are prepared with relatively nontoxicacids or bases, depending on the particular substituents found on thecompounds described herein. When compounds of the present inventioncontain relatively acidic functionalities, base addition salts can beobtained by contacting the neutral form of such compounds with asufficient amount of the desired base, either neat or in a suitableinert solvent. Examples of salts derived frompharmaceutically-acceptable inorganic bases include aluminum, ammonium,calcium, copper, ferric, ferrous, lithium, magnesium, manganic,manganous, potassium, sodium, zinc and the like. Salts derived frompharmaceutically-acceptable organic bases include salts of primary,secondary and tertiary amines, including substituted amines, cyclicamines, naturally-occurring amines and the like, such as arginine,betaine, caffeine, choline, N,N′-dibenzylethylenediamine, diethylamine,2-diethylaminoethanol, 2-dimethylaminoethanol, ethanolamine,ethylenediamine, N-ethylmorpholine, N-ethylpiperidine, glucamine,glucosamine, histidine, hydrabamine, isopropylamine, lysine,methylglucamine, morpholine, piperazine, piperadine, polyamine resins,procaine, purines, theobromine, triethylamine, trimethylamine,tripropylamine, tromethamine and the like. When compounds of the presentinvention contain relatively basic functionalities, acid addition saltscan be obtained by contacting the neutral form of such compounds with asufficient amount of the desired acid, either neat or in a suitableinert solvent. Examples of pharmaceutically acceptable acid additionsalts include those derived from inorganic acids like hydrochloric,hydrobromic, nitric, carbonic, monohydrogencarbonic, phosphoric,monohydrogenphosphoric, dihydrogenphosphoric, sulfuric,monohydrogensulfuric, hydriodic, or phosphorous acids and the like, aswell as the salts derived from relatively nontoxic organic acids likeacetic, propionic, isobutyric, malonic, benzoic, succinic, suberic,fumaric, mandelic, phthalic, benzenesulfonic, p-tolylsulfonic, citric,tartaric, methanesulfonic, and the like. Also included are salts ofamino acids such as arginate and the like, and salts of organic acidslike glucuronic or galactunoric acids and the like (see, for example,Berge, S. M., et al, “Pharmaceutical Salts”, Journal of PharmaceuticalScience, 1977, 66, 1-19). Certain specific compounds of the presentinvention contain both basic and acidic functionalities that allow thecompounds to be converted into either base or acid addition salts.

The neutral forms of the compounds may be regenerated by contacting thesalt with a base or acid and isolating the parent compound in theconventional manner. The parent form of the compound differs from thevarious salt forms in certain physical properties, such as solubility inpolar solvents, but otherwise the salts are equivalent to the parentform of the compound for the purposes of the present invention.

In addition to salt forms, the present invention provides compoundswhich are in a prodrug form. Prodrugs of the compounds described hereinare those compounds that readily undergo chemical changes underphysiological conditions to provide the compounds of the presentinvention. Additionally, prodrugs can be converted to the compounds ofthe present invention by chemical or biochemical methods in an ex vivoenvironment. For example, prodrugs can be slowly converted to thecompounds of the present invention when placed in a transdermal patchreservoir with a suitable enzyme or chemical reagent.

Certain compounds of the present invention can exist in unsolvated formsas well as solvated forms, including hydrated forms. In general, thesolvated forms are equivalent to unsolvated forms and are intended to beencompassed within the scope of the present invention. Certain compoundsof the present invention may exist in multiple crystalline or amorphousforms. In general, all physical forms are equivalent for the usescontemplated by the present invention and are intended to be within thescope of the present invention.

Certain compounds of the present invention possess asymmetric carbonatoms (optical centers) or double bonds; the racemates, diastereomers,geometric isomers, regioisomers and individual isomers (e.g., separateenantiomers) are all intended to be encompassed within the scope of thepresent invention. The compounds of the present invention may alsocontain unnatural proportions of atomic isotopes at one or more of theatoms that constitute such compounds. For example, the compounds may beradiolabeled with radioactive isotopes, such as for example tritium(³H), iodine-125 (¹²⁵I) or carbon-14 (¹⁴C). All isotopic variations ofthe compounds of the present invention, whether radioactive or not, areintended to be encompassed within the scope of the present invention.

As used herein, a wavy line, “

”, that intersects a single, double or triple bond in any chemicalstructure depicted herein, represent the point attachment of the single,double, or triple bond to the remainder of the molecule.

II. Description of the Embodiments

A. Compounds

In one aspect, the present invention provides compounds of Formula (I):

-   -   or a pharmaceutically acceptable salt thereof, wherein,    -   ring vertex a is N or C(R^(2c)), ring vertex b is N or        C(R^(2d)), and ring vertex e is N or C(R^(2e)), wherein no more        than one of a, b and e is N;    -   X¹ is selected from the group consisting of a bond, C₁₋₈        alkylene, C(O), C(O)—C₁₋₄ alkylene, and S(O)₂;    -   R¹ is selected from the group consisting of        -   a) 5- to 10-membered heteroaryl having from 1 to 4            heteroatoms as ring vertices selected from N, O and S;        -   b) C₆₋₁₀ aryl;        -   c) C₃₋₈ cycloalkyl;        -   d) 4- to 8-membered heterocycloalkyl having from 1 to 2            heteroatoms as ring vertices selected from N, O and S; and        -   e) C₁₋₈ alkyl, C₁₋₈ alkoxy, C₁₋₈ haloalkyl,            —C(O)NR^(1a)R^(1b), and —CO₂R^(1a); wherein R^(1a) and            R^(1b) are each independently selected from the group            consisting of hydrogen, C₁₋₈ alkyl, C₆₋₁₀ aryl, and —C₁₋₆            alkylene-C₆₋₁₀ aryl;    -   wherein the group —X¹—R¹ is optionally substituted with 1 to 5        R^(x) substituents;    -   R^(2a) and R^(2e) are each independently selected from the group        consisting of hydrogen, C₁₋₆ alkyl, C₁₋₆ alkoxy, C₁₋₆ haloalkyl,        —O—C₁₋₆ haloalkyl, —S—C₁₋₆ alkyl, —C₁₋₆ alkyl-O—C₁₋₆ alkyl,        —C₁₋₆ alkyl-S—C₁₋₆ alkyl, CN, and halogen, and at least one of        R^(2a) and R^(2e) is other than hydrogen;    -   R^(2b), R^(2c), and R^(2d) are each independently selected from        the group consisting of hydrogen, C₁₋₆ alkyl, C₁₋₆ alkoxy, C₁₋₆        haloalkyl, —O—C₁₋₆ haloalkyl, —S—C₁₋₆ alkyl, —C₁₋₆ alkyl-O—C₁₋₆        alkyl, —C₁₋₆ alkyl-S—C₁₋₆ alkyl, cyano, and halogen;    -   each R³ is independently selected from the group consisting of        hydroxyl, C₁₋₄ alkyl, C₁₋₄ haloalkyl and C₁₋₄ hydroxyalkyl, and        optionally two R³ groups on the same carbon atom are combined to        form oxo (═O), and optionally two R³ groups and the carbon atoms        they are attached to form a 3-6 membered ring with 0-2        hetereoatoms as ring members selected from O, N, and S;    -   R⁴ is independently selected from the group consisting of        —X²—OR^(4a), —X²—NR^(4a)R^(4b), —X²—CONR^(4a)R^(4b),        —X²—NR^(4a)—C(O)NR^(4a)R^(4b), —X²—NR^(4a)—C(O)—C₁₋₃        alkylene-OR^(4a) and —X²—NR^(4a)—C(O)—C₁₋₃        alkylene-NR^(4a)R^(4b); wherein each X² is independently a bond,        C(O), C₁₋₄ alkylene, C(O)—C₁₋₄ alkylene, and C₁₋₄ alkylene-C(O),        and each R^(4a) and R^(4b) is independently selected from the        group consisting of hydrogen, C₁₋₄ alkyl, and C₁₋₄ haloalkyl;    -   each R⁵ is independently selected from the group consisting of        C₁₋₈ alkyl, C₁₋₈ alkoxy, C₁₋₈ haloalkyl, C₁₋₈ haloalkoxy, C₁₋₈        hydroxyalkyl, halogen, OH, CN, C(O)R^(5a) and CO₂R^(5a); wherein        each R^(5a) is independently selected from the group consisting        of hydrogen, C₁₋₄ alkyl, and C₁₋₄ haloalkyl;    -   each R^(x) is independently selected from the group consisting        of halogen, CN, C₁₋₄ alkyl, C₁₋₄ alkoxy, C₁₋₄ haloalkyl, C₁₋₄        haloalkoxy, C₁₋₄ hydroxy, C₂₋₄ alkenyl, C₃₋₆ cycloalkyl,        CO₂—C₁₋₄ alkyl, and CONH₂;    -   the subscript m is 0, 1, 2, 3 or 4; and    -   the subscript n is 0, 1, 2 or 3.

In some embodiments, the compounds of formula (I) are represented byformula (Ia) or (Ib):

In one group of embodiments for the compounds of formula (I), (Ia) or(Ib), R⁴ is selected from the group consisting of

In another group of embodiments for the compounds of formula (I), (Ia)or (Ib), or a pharmaceutically acceptable salt thereof, R⁴ is selectedfrom the group consisting of

In yet another group of embodiments for the compounds of formula (I),(Ia) or (Ib), or a pharmaceutically acceptable salt thereof, wherein R⁴is selected from the group consisting of

With reference to the compounds of formula (I), (Ia) or (Ib), or apharmaceutically acceptable salt thereof, as well as any of the groupsof embodiments noted above, in certain selected embodiments, X¹ is abond; in other selected embodiments, X¹ is C(O); in still other selectedembodiments, X¹ is C₁₋₈ alkylene; in yet other selected embodiments, X¹is C(O)—C₁₋₄ alkylene or S(O)₂.

With reference to the compounds of formula (I), (Ia) or (Ib), or apharmaceutically acceptable salt thereof, as well as any of the groupsor selected embodiments noted above, in some further embodiments,wherein R¹ is a 5- to 10-membered heteroaryl having from 1 to 4heteroatoms as ring vertices selected from N, O and S; and wherein thegroup —X¹—R¹ is optionally substituted with 1 to 4 R^(x) substituents.In still further embodiments, R¹ is selected from the group consistingof pyrazolyl, pyridyl, pyrimidinyl, imidazolyl, thiazolyl, thiadiazolyland pyrazinyl; and wherein the group —X¹—R¹ is optionally substitutedwith 1 to 4 R^(x) substituents.

With reference to the compounds of formula (I), (Ia) or (Ib), or apharmaceutically acceptable salt thereof, as well as any of the groupsor selected embodiments noted above, in some further embodiments,wherein R¹ is C₆₋₁₀ aryl; and wherein the group —X¹—R¹ is optionallysubstituted with 1 to 4 R^(x) substituents. In still furtherembodiments, R¹ is phenyl; and wherein the group —X¹—R¹ is optionallysubstituted with 1 to 4 R^(x) substituents.

With reference to the compounds of formula (I), (Ia) or (Ib), or apharmaceutically acceptable salt thereof, as well as any of the groupsor selected embodiments noted above, in some further embodiments, R¹ isC₃₋₈ cycloalkyl; and wherein the group —X¹—R¹ is optionally substitutedwith 1 to 4 R^(x) substituents. In still further embodiments, R¹ isselected from the group consisting of cyclobutyl, cyclopentyl andcyclohexyl; and wherein the group —X¹—R¹ is optionally substituted with1 to 4 R^(x) substituents.

With reference to the compounds of formula (I), (Ia) or (Ib), or apharmaceutically acceptable salt thereof, as well as any of the groupsor selected embodiments noted above, in some further embodiments, R¹ isa 4- to 8-membered heterocycloalkyl having from 1 to 2 heteroatoms asring vertices selected from N, O and S; and wherein the group —X¹—R¹ isoptionally substituted with 1 to 4 R^(x) substituents. In still furtherselected embodiments, R¹ is selected from the group consisting ofoxetanyl, tetrahydrofuranyl, tetrahydropyranyl and morpholinyl; andwherein the group —X¹—R¹ is optionally substituted with 1 to 4 R^(x)substituents.

With reference to the compounds of formula (I), (Ia) or (Ib), or apharmaceutically acceptable salt thereof, as well as any of the groupsor selected embodiments noted above, in some further embodiments, R¹ isselected from the group consisting of C₁₋₈ alkyl, C₁₋₈ alkoxy, C₁₋₈haloalkyl, —C(O)NR^(1a)R^(1b), and —CO₂R^(1a); wherein R^(1a) and R^(1b)are each independently selected from the group consisting of hydrogen,C₁₋₈ alkyl, C₆₋₁₀ aryl, and —C₁₋₆ alkylene-C₆₋₁₀ aryl; and wherein thegroup —X¹—R¹ is optionally substituted with 1 to 4 R^(x) substituents.

With reference to the compounds of formula (I), (Ia) or (Ib), or apharmaceutically acceptable salt thereof, as well as any of the groupsor selected embodiments noted above, in some further embodiments, R¹ isselected from the group consisting of phenyl, pyridyl, pyrimidinyl, andpyrazinyl; and wherein the group —X¹—R¹ is optionally substituted with 1to 4 R^(x) substituents.

With reference to the compounds of formula (I), or a pharmaceuticallyacceptable salt thereof, as well as any of the embodiments noted above,in some further embodiments, ring vertices a and b are CH; R^(2b) is H;ring vertex e is C(R^(2e)), and R^(2a) and R^(2e) are independentlyselected from the group consisting of C₁₋₆ alkyl, C₁₋₆ alkoxy, C₁₋₆haloalkyl, —O—C₁₋₆ haloalkyl, —S—C₁₋₆ alkyl, —C₁₋₆ alkyl-O—C₁₋₆ alkyl,—C₁₋₆ alkyl-S—C₁₋₆ alkyl, CN, and halogen.

With reference to the compounds of formula (I), or a pharmaceuticallyacceptable salt thereof, as well as any of the embodiments noted above,in some further embodiments, ring vertices a and b are CH; R^(2b) is H;ring vertex e is C(R^(2e)), and R^(2a) and R^(2e) are independentlyselected from the group consisting of C₁₋₆ alkyl, C₁₋₆ alkoxy andhalogen.

With reference to the compounds of formula (I), (Ia) or (Ib), or apharmaceutically acceptable salt thereof, as well as any of theembodiments noted above, in some further embodiments, the subscript n is0, 1 or 2 and each R⁵, when present, is selected from the groupconsisting of F, Cl, CN, C₁₋₄ alkyl and C₁₋₄ alkoxy. In still furtherselected embodiments, the subscript n is 0, 1 or 2 and each R⁵, whenpresent, is selected from the group consisting of F, Cl, CN, CH₃ andOCH₃.

With reference to the compounds of formula (I), or a pharmaceuticallyacceptable salt thereof, as well as any of the embodiments noted above,in some further embodiments, the subscript m is 0, 1 or 2 and each R³,when present, is C₁₋₄ alkyl.

In a particular group of embodiments of the compounds of formula (I), ora pharmaceutically acceptable salt thereof, R¹ is selected from thegroup consisting of phenyl or pyridyl, wherein the group —X¹—R¹ isoptionally substituted with 1 to 4 R^(x) substituents; ring vertices aand b are CH; R^(2b) is H; ring vertex e is C(R^(2e)), and R^(2a) andR^(2e) are independently selected from the group consisting of C₁₋₆alkyl, C₁₋₆ alkoxy and halogen; m is 0, 1 or 2 and each R³, whenpresent, is CH₃; R⁴ is selected from the group consisting of

n is 0, 1 or 2 and each R⁵, when present, is selected from the groupconsisting of F, Cl, CN, CH₃ and OCH₃.

In some embodiments of the compounds of formula (I), or apharmaceutically acceptable salt thereof, —X¹—R¹ is selected from thegroup consisting of:

With reference to the compounds of formula (I), or a pharmaceuticallyacceptable salt thereof, as well as any of the embodiments noted above,in some further embodiments, the group

a is selected from the group consisting of

With reference to the compounds of formula (I), or a pharmaceuticallyacceptable salt thereof, as well as any of the embodiments noted above,in some further embodiments, wherein n is 0.

With reference to the compounds of formula (I), or a pharmaceuticallyacceptable salt thereof, as well as any of the embodiments noted above,in some further embodiments, the subscript n is 2 and the two R³ groupsare on the same carbon atom are each methyl, or are combined to form oxo(═O).

In some selected embodiments, provided herein is a compound selectedfrom the group consisting of:

or a pharmaceutically acceptable salt thereof.

In some embodiments, the compound of Formula (I) is a compound describedin the Examples section and the accompanying Tables.

Preparation of Compounds

Certain compounds of the invention can be prepared following methodologyas described in the Examples section of this document. In addition, thesyntheses of certain intermediate compounds that are useful in thepreparation of compounds of the invention are also described.

B. Pharmaceutical Compositions

In addition to the compounds provided above, compositions for modulatingC5a activity in humans and animals will typically contain apharmaceutical carrier or diluent.

The term “composition” as used herein is intended to encompass a productcomprising the specified ingredients in the specified amounts, as wellas any product which results, directly or indirectly, from combinationof the specified ingredients in the specified amounts. By“pharmaceutically acceptable” it is meant the carrier, diluent orexcipient must be compatible with the other ingredients of theformulation and not deleterious to the recipient thereof.

The pharmaceutical compositions for the administration of the compoundsof this invention may conveniently be presented in unit dosage form andmay be prepared by any of the methods well known in the art of pharmacyand drug delivery. All methods include the step of bringing the activeingredient into association with the carrier which constitutes one ormore accessory ingredients. In general, the pharmaceutical compositionsare prepared by uniformly and intimately bringing the active ingredientinto association with a liquid carrier or a finely divided solid carrieror both, and then, if necessary, shaping the product into the desiredformulation. In the pharmaceutical composition the active objectcompound is included in an amount sufficient to produce the desiredeffect upon the process or condition of diseases.

The pharmaceutical compositions containing the active ingredient may bein a form suitable for oral use, for example, as tablets, troches,lozenges, aqueous or oily suspensions, dispersible powders or granules,emulsions and self emulsifications as described in U.S. PatentApplication 2002-0012680, hard or soft capsules, syrups, elixirs,solutions, buccal patch, oral gel, chewing gum, chewable tablets,effervescent powder and effervescent tablets. Compositions intended fororal use may be prepared according to any method known to the art forthe manufacture of pharmaceutical compositions and such compositions maycontain one or more agents selected from the group consisting ofsweetening agents, flavoring agents, coloring agents, antioxidants andpreserving agents in order to provide pharmaceutically elegant andpalatable preparations. Tablets contain the active ingredient inadmixture with non-toxic pharmaceutically acceptable excipients whichare suitable for the manufacture of tablets. These excipients may be forexample, inert diluents, such as cellulose, silicon dioxide, aluminumoxide, calcium carbonate, sodium carbonate, glucose, mannitol, sorbitol,lactose, calcium phosphate or sodium phosphate; granulating anddisintegrating agents, for example, corn starch, or alginic acid;binding agents, for example PVP, cellulose, PEG, starch, gelatin oracacia, and lubricating agents, for example magnesium stearate, stearicacid or talc. The tablets may be uncoated or they may be coated,enterically or otherwise, by known techniques to delay disintegrationand absorption in the gastrointestinal tract and thereby provide asustained action over a longer period. For example, a time delaymaterial such as glyceryl monostearate or glyceryl distearate may beemployed. They may also be coated by the techniques described in theU.S. Pat. Nos. 4,256,108; 4,166,452; and 4,265,874 to form osmotictherapeutic tablets for control release.

Formulations for oral use may also be presented as hard gelatin capsuleswherein the active ingredient is mixed with an inert solid diluent, forexample, calcium carbonate, calcium phosphate or kaolin, or as softgelatin capsules wherein the active ingredient is mixed with water or anoil medium, for example peanut oil, liquid paraffin, or olive oil.Additionally, emulsions can be prepared with a non-water miscibleingredient such as oils and stabilized with surfactants such asmono-diglycerides, PEG esters and the like.

Aqueous suspensions contain the active materials in admixture withexcipients suitable for the manufacture of aqueous suspensions. Suchexcipients are suspending agents, for example sodiumcarboxymethylcellulose, methylcellulose, hydroxy-propylmethylcellulose,sodium alginate, polyvinyl-pyrrolidone, gum tragacanth and gum acacia;dispersing or wetting agents may be a naturally-occurring phosphatide,for example lecithin, or condensation products of an alkylene oxide withfatty acids, for example polyoxy-ethylene stearate, or condensationproducts of ethylene oxide with long chain aliphatic alcohols, forexample heptadecaethyleneoxycetanol, or condensation products ofethylene oxide with partial esters derived from fatty acids and ahexitol such as polyoxyethylene sorbitol monooleate, or condensationproducts of ethylene oxide with partial esters derived from fatty acidsand hexitol anhydrides, for example polyethylene sorbitan monooleate.The aqueous suspensions may also contain one or more preservatives, forexample ethyl, or n-propyl, p-hydroxybenzoate, one or more coloringagents, one or more flavoring agents, and one or more sweetening agents,such as sucrose or saccharin.

Oily suspensions may be formulated by suspending the active ingredientin a vegetable oil, for example arachis oil, olive oil, sesame oil orcoconut oil, or in a mineral oil such as liquid paraffin. The oilysuspensions may contain a thickening agent, for example beeswax, hardparaffin or cetyl alcohol. Sweetening agents such as those set forthabove, and flavoring agents may be added to provide a palatable oralpreparation. These compositions may be preserved by the addition of ananti-oxidant such as ascorbic acid.

Dispersible powders and granules suitable for preparation of an aqueoussuspension by the addition of water provide the active ingredient inadmixture with a dispersing or wetting agent, suspending agent and oneor more preservatives. Suitable dispersing or wetting agents andsuspending agents are exemplified by those already mentioned above.Additional excipients, for example sweetening, flavoring and coloringagents, may also be present.

The pharmaceutical compositions of the invention may also be in the formof oil-in-water emulsions. The oily phase may be a vegetable oil, forexample olive oil or arachis oil, or a mineral oil, for example liquidparaffin or mixtures of these. Suitable emulsifying agents may benaturally-occurring gums, for example gum acacia or gum tragacanth,naturally-occurring phosphatides, for example soy bean, lecithin, andesters or partial esters derived from fatty acids and hexitolanhydrides, for example sorbitan monooleate, and condensation productsof the said partial esters with ethylene oxide, for examplepolyoxyethylene sorbitan monooleate. The emulsions may also containsweetening and flavoring agents.

Syrups and elixirs may be formulated with sweetening agents, for exampleglycerol, propylene glycol, sorbitol or sucrose. Such formulations mayalso contain a demulcent, a preservative and flavoring and coloringagents. Oral solutions can be prepared in combination with, for example,cyclodextrin, PEG and surfactants.

The pharmaceutical compositions may be in the form of a sterileinjectable aqueous or oleagenous suspension. This suspension may beformulated according to the known art using those suitable dispersing orwetting agents and suspending agents which have been mentioned above.The sterile injectable preparation may also be a sterile injectablesolution or suspension in a non-toxic parenterally-acceptable diluent orsolvent, for example as a solution in 1,3-butane diol. Among theacceptable vehicles and solvents that may be employed are water,Ringer's solution and isotonic sodium chloride solution. In addition,sterile, fixed oils are conventionally employed as a solvent orsuspending medium. For this purpose any bland fixed oil may be employedincluding synthetic mono- or diglycerides. In addition, fatty acids suchas oleic acid find use in the preparation of injectables.

The compounds of the present invention may also be administered in theform of suppositories for rectal administration of the drug. Thesecompositions can be prepared by mixing the drug with a suitablenon-irritating excipient which is solid at ordinary temperatures butliquid at the rectal temperature and will therefore melt in the rectumto release the drug. Such materials include cocoa butter andpolyethylene glycols. Additionally, the compounds can be administeredvia ocular delivery by means of solutions or ointments. Still further,transdermal delivery of the subject compounds can be accomplished bymeans of iontophoretic patches and the like. For topical use, creams,ointments, jellies, solutions or suspensions, etc., containing thecompounds of the present invention are employed. As used herein, topicalapplication is also meant to include the use of mouth washes andgargles.

The compounds of this invention may also be coupled a carrier that is asuitable polymers as targetable drug carriers. Such polymers can includepolyvinylpyrrolidone, pyran copolymer,polyhydroxy-propyl-methacrylamide-phenol,polyhydroxyethyl-aspartamide-phenol, or polyethyleneoxide-polylysinesubstituted with palmitoyl residues. Furthermore, the compounds of theinvention may be coupled to a carrier that is a class of biodegradablepolymers useful in achieving controlled release of a drug, for examplepolylactic acid, polyglycolic acid, copolymers of polylactic andpolyglycolic acid, polyepsilon caprolactone, polyhydroxy butyric acid,polyorthoesters, polyacetals, polydihydropyrans, polycyanoacrylates andcross linked or amphipathic block copolymers of hydrogels. Polymers andsemipermeable polymer matrices may be formed into shaped articles, suchas valves, stents, tubing, prostheses and the like. In one embodiment ofthe invention, the compound of the invention is coupled to a polymer orsemipermeable polymer matrix that is formed as a stent or stent-graftdevice.

The pharmaceutical compositions of the present disclosure may beformulated with one or more additional therapeutic agents. The one ormore additional therapeutic agent is selected from the group consistingof corticosteroids, steroids, immunosuppressants, Immunoglobulin Gagonists, Dipeptidyl peptidase IV inhibitors, Lymphocyte functionantigen-3 receptor antagonists, Interleukin-2 ligands, Interleukin-1beta ligand inhibitors, IL-2 receptor alpha subunit inhibitors, HGF genestimulators, IL-6 antagonists, IL-5 antagonists, Alpha 1 antitrypsinstimulators, Cannabinoid receptor antagonists, Histone deacetylaseinhibitors, AKT protein kinase inhibitors, CD20 inhibitors, Abl tyrosinekinase inhibitors, JAK tyrosine kinase inhibitors, TNF alpha ligandinhibitors, Hemoglobin modulators, TNF antagonists, proteasomeinhibitors, CD3 modulators, Hsp 70 family inhibitors, Immunoglobulinagonists, CD30 antagonists, tubulin antagonists, Sphingosine-1-phosphatereceptor-1 agonists, connective tissue growth factor ligand inhibitors,caspase inhibitors, adrenocorticotrophic hormone ligands, Btk tyrosinekinase inhibitors, Complement C is subcomponent inhibitors,Erythropoietin receptor agonists, B-lymphocyte stimulator ligandinhibitors, Cyclin-dependent kinase-2 inhibitors, P-selectinglycoprotein ligand-1 stimulators, mTOR inhibitors, Elongation factor 2inhibitors, Cell adhesion molecule inhibitors, Factor XIII agonists,Calcineurin inhibitors, Immunoglobulin G1 agonists, Inosinemonophosphate dehydrogenase inhibitors, Complement C is subcomponentinhibitors, Thymidine kinase modulators, Cytotoxic T-lymphocyteprotein-4 modulators, Angiotensin II receptor antagonists, AngiotensinII receptor modulators, TNF superfamily receptor 12A antagonists, CD52antagonists, Adenosine deaminase inhibitors, T-cell differentiationantigen CD6 inhibitors, FGF-7 ligands, dihydroorotate dehydrogenaseinhibitors, Syk tyrosine kinase inhibitors, Interferon type I receptorantagonists, Interferon alpha ligand inhibitors, Macrophage migrationinhibitory factor inhibitors, Integrin alpha-V/beta-6 antagonists,Cysteine protease stimulators, p38 MAP kinase inhibitors, TP53 geneinhibitors, Shiga like toxin I inhibitors, Fucosyltransferase 6stimulators, Interleukin 22 ligands, IRS1 gene inhibitors, Proteinkinase C stimulators, Protein kinase C alpha inhibitors, CD74antagonists, Immunoglobulin gamma Fc receptor IIB antagonists, T-cellantigen CD7 inhibitors, CD95 antagonists, N acetylmannosamine kinasestimulators, Cardiotrophin-1 ligands, Leukocyte elastase inhibitors,CD40 ligand receptor antagonists, CD40 ligand modulators, IL-17antagonists, TLR-2 antagonists, Mannan-binding lectin serine protease-2(MASP-2) inhibitors, Factor B inhibitors, Factor D inhibitors, C3aRmodulators, C5aR2 modulators, T cell receptor antagonists, PD-1inhibitors, PD-L1 inhibitors, TIGIT inhibitors, TIM-3 inhibitors, LAG-3inhibitors, VISTA inhibitors, STING agonists, IDO inhibitors, adenosinereceptor modulators, CD39 inhibitors, CD73 inhibitors, antagonists ofthe chemokine receptors, especially CXCR1, CXCR2, CXCR3, CXCR4, CXCR7,CCR1, CCR2, CCR3, CCR4, CCR5, CCR7, CCR7, CCR9, CX3CR1 and CXCR6, andcombinations thereof.

In some embodiments, the one or more additional therapeutic agent isselected from the group consisting of obinutuzumab, rituximab,ocrelizumab, cyclophosphamide, prednisone, hydrocortisone,hydrocortisone acetate, cortisone acetate, tixocortol pivalate,prednisolone, methylprednisolone, triamcinolone acetonide, triamcinolonealcohol, mometasone, amcinonide, budesonide, desonide, fluocinonide,fluocinolone acetonide, halcinonide, betamethasone, betamethasone sodiumphosphate, dexamethasone, dexamethasone sodium phosphate, fluocortolone,hydrocortisone-17-valerate, halometasone, alclometasone dipropionate,beclomethasone, betamethasone valerate, betamethasone dipropionate,prednicarbate, clobetasone-17-butyrate, clobetasol-17-propionate,fluocortolone caproate, fluocortolone pivalate, fluprednidene acetate,hydrocortisone-17-butyrate, hydrocortisone-17-aceponate,hydrocortisone-17-buteprate, ciclesonide and prednicarbate, GB-0998,immuglo, begelomab, alefacept, aldesleukin, gevokizumab, daclizumab,basiliximab, inolimomab, beperminogene perplasmid, sirukumab,tocilizumab, clazakizumab, mepolizumab, fingolimod, panobinostat,triciribine, nilotinib, imatinib, tofacitinib, momelotinib, peficitinib,itacitinib, infliximab, PEG-bHb-CO, etanercept, ixazomib, bortezomib,muromonab, otelixizumab, gusperimus, brentuximab vedotin, Ponesimod,KRP-203, FG-3019, emricasan, corticotropin, ibrutinib, cinryze,conestat, methoxy polyethylene glycol-epoetin beta, belimumab,blisibimod, atacicept, seliciclib, neihulizumab, everolimus, sirolimus,denileukin diftitox, LMB-2, natalizumab, catridecacog, ciclosporin,tacrolimus, voclosporin, voclosporin, canakinumab, mycophenolate,mizoribine, CE-1145, TK-DLI, abatacept, belatacept, olmesartanmedoxomil, sparsentan, TXA-127, BIIB-023, alemtuzumab, pentostatin,itolizumab, palifermin, leflunomide, PRO-140, cenicriviroc,fostamatinib, anifrolumab, sifalimumab, BAX-069, BG-00011, losmapimod,QPI-1002, ShigamAbs, TZ-101, F-652, reparixin, ladarixin, PTX-9908,aganirsen, APH-703, sotrastaurin, sotrastaurin, milatuzumab, SM-101,T-Guard, APG-101, DEX-M74, cardiotrophin-1, tiprelestat, ASKP-1240,BMS-986004, HPH-116, KD-025, OPN-305, TOL-101, defibrotide,pomalidomide, Thymoglobulin, laquinimod, remestemcel-L, Equineantithymocyte immunoglobulin, Stempeucel, LIV-Gamma, Octagam 10%,t2c-001, 99mTc-sestamibi, Clairyg, Prosorba, pomalidomide, laquinimod,teplizumab, FCRx, solnatide, foralumab, ATIR-101, BPX-501, ACP-01,ALLO-ASC-DFU, irbesartan+propagermanium, ApoCell, cannabidiol, RGI-2001,saratin, anti-CD3 bivalent antibody-diphtheria toxin conjugate, NOX-100,LT-1951, OMS721, ALN-CC5, ACH-4471, AMY-101, Acthar gel, and CD4+CD25+regulatory T-cells, MEDI7814, P32, P59, pembrolizumab, nivolumab,atezolizumab, avelumab, durvalumab, CCX354, CCX721, CCX9588, CCX140,CCX872, CCX598, CCX6239, CCX587, CCX624, CCX282, CCX025, CCX507, CCX430,CCX765, CCX758, CCX771, CCX662, CCX650, and combinations thereof.Further discussions of combination therapy are included in the “Methodsof Use” section of this application.

C. Methods of Use

The compounds of the invention may be used as agonists, (preferably)antagonists, partial agonists, inverse agonists, of C5a receptors in avariety of contexts, both in vitro and in vivo. In one embodiment, thecompounds of the invention are C5aR antagonist that can be used toinhibit the binding of C5a receptor ligand (e.g., C5a) to C5a receptorin vitro or in vivo. In general, such methods comprise the step ofcontacting a C5a receptor with a sufficient amount of one or more C5areceptor modulators as provided herein, in the presence of C5a receptorligand in aqueous solution and under conditions otherwise suitable forbinding of the ligand to C5a receptor. The C5a receptor may be presentin suspension (e.g., in an isolated membrane or cell preparation), in acultured or isolated cell, or in a tissue or organ.

Preferably, the amount of C5a receptor modulator contacted with thereceptor should be sufficient to inhibit C5a binding to C5a receptor invitro as measured, for example, using a radioligand binding assay,calcium mobilization assay, or chemotaxis assay as described herein.

In one embodiment of the invention, the C5a modulators of the inventionare used to modulate, preferably inhibit, the signal-transducingactivity of a C5a receptor, for example, by contacting one or morecompound(s) of the invention with a C5a receptor (either in vitro or invivo) under conditions suitable for binding of the modulator(s) to thereceptor. The receptor may be present in solution or suspension, in acultured or isolated cell preparation or within a patient. Anymodulation of the signal transducing activity may be assessed bydetecting an effect on calcium ion calcium mobilization or by detectingan effect on C5a receptor-mediated cellular chemotaxis. In general, aneffective amount of C5a modulator(s) is an amount sufficient to modulateC5a receptor signal transducing activity in vitro within a calciummobilization assay or C5a receptor-mediated cellular chemotaxis within amigration assay.

When compounds of the invention are used to inhibit C5areceptor-mediated cellular chemotaxis, preferably leukocyte (e.g.,neutrophil) chemotaxis, in an in vitro chemotaxis assay, such methodscomprise contacting white blood cells (particularly primate white bloodcells, especially human white blood cells) with one or more compounds ofthe invention. Preferably the concentration is sufficient to inhibitchemotaxis of white blood cells in an in vitro chemotaxis assay, so thatthe levels of chemotaxis observed in a control assay are significantlyhigher, as described above, than the levels observed in an assay towhich a compound of the invention has been added.

In another embodiment, the compounds of the present invention furthercan be used for treating patients suffering from conditions that areresponsive to C5a receptor modulation. As used herein, the term“treating” or “treatment” encompasses both disease-modifying treatmentand symptomatic treatment, either of which may be prophylactic (i.e.,before the onset of symptoms, in order to prevent, delay or reduce theseverity of symptoms) or therapeutic (i.e., after the onset of symptoms,in order to reduce the severity and/or duration of symptoms). As usedherein, a condition is considered “responsive to C5a receptormodulation” if modulation of C5a receptor activity results in thereduction of inappropriate activity of a C5a receptor. As used herein,the term “patients” include primates (especially humans), domesticatedcompanion animals (such as dogs, cats, horses, and the like) andlivestock (such as cattle, pigs, sheep, and the like), with dosages asdescribed herein.

Conditions that can be Treated by C5a Modulation:

Autoimmune disorders—e.g., Rheumatoid arthritis, systemic lupuserythematosus, Guillain-Barre syndrome, pancreatitis, lupus nephritis,lupus glomerulonephritis, psoriasis, Crohn's disease, vasculitis,irritable bowel syndrome, dermatomyositis, multiple sclerosis, bronchialasthma, dense deposit disease, pemphigus, pemphigoid, scleroderma,myasthenia gravis, autoimmune hemolytic and thrombocytopenic states,Goodpasture's syndrome (and associated glomerulonephritis and pulmonaryhemorrhage), C3-glomerulopathy, C3-glomerulonephritis,membranoproliferative glomerulonephritis, Kawasaki disease, IGsnephropathy, immunovasculitis, tissue graft rejection, graft versus hostdisease, hyperacute rejection of transplanted organs; and the like.

Inflammatory disorders and related conditions—e.g., Neutropenia, sepsis,septic shock, Alzheimer's disease, multiple sclerosis, neutrophilia,stroke, inflammatory bowel disease (IBD), inflammation associated withsevere burns, lung injury, and ischemia-reperfusion injury,osteoarthritis, as well as acute (adult) respiratory distress syndrome(ARDS), chronic pulmonary obstructive disorder (COPD), systemicinflammatory response syndrome (SIRS), atopic dermatitis, psoriasis,chronic urticaria and multiple organ dysfunction syndrome (MODS)Hemolytic uremic syndrome, atypical hemolytic uremic syndrome (aHUS).Also included are pathologic sequellae associated with insulin-dependentdiabetes mellitus (including diabetic retinopathy), lupus nephropathy,Heyman nephritis, membranous nephritis and other forms ofglomerulonephritis, contact sensitivity responses, and inflammationresulting from contact of blood with artificial surfaces that can causecomplement activation, as occurs, for example, during extracorporealcirculation of blood (e.g., during hemodialysis or via a heart-lungmachine, for example, in association with vascular surgery such ascoronary artery bypass grafting or heart valve replacement), or inassociation with contact with other artificial vessel or containersurfaces (e.g., ventricular assist devices, artificial heart machines,transfusion tubing, blood storage bags, plasmapheresis,plateletpheresis, and the like). Also included are diseases related toischemia/reperfusion injury, such as those resulting from transplants,including solid organ transplant, and syndromes such as ischemicreperfusion injury, ischemic colitis and cardiac ischemia. Compounds ofthe instant invention may also be useful in the treatment of age-relatedmacular degeneration (Hageman et al, P.N.A.S. 102: 7227-7232, 2005).

Cardiovascular and Cerebrovascular Disorders—e.g., myocardialinfarction, coronary thrombosis, vascular occlusion, post-surgicalvascular reocclusion, atherosclerosis, traumatic central nervous systeminjury, and ischemic heart disease. In one embodiment, an effectiveamount of a compound of the invention may be administered to a patientat risk for myocardial infarction or thrombosis (i.e., a patient who hasone or more recognized risk factor for myocardial infarction orthrombosis, such as, but not limited to, obesity, smoking, high bloodpressure, hypercholesterolemia, previous or genetic history ofmyocardial infarction or thrombosis) in order reduce the risk ofmyocardial infarction or thrombosis.

Oncologic Diseases or Disorders—e.g., melanoma, lung cancer, lymphoma,sarcoma, carcinoma, fibrosarcoma, liposarcoma, chondrosarcoma,osteogenic sarcoma, angiosarcoma, lymphangiosarcoma, synovioma,mesothelioma, meningioma, leukemia, lymphoma, leiomyosarcoma,rhabdomyosarcoma, squamous cell carcinoma, basal cell carcinoma,adenocarcinoma, papillary carcinoma, cystadenocarcinoma, bronchogeniccarcinoma, renal cell carcinoma, hepatocellular carcinoma, transitionalcell carcinoma, choriocarcinoma, seminoma, embryonal carcinoma, wilm'stumor, pleomorphic adenoma, liver cell papilloma, renal tubular adenoma,cystadenoma, papilloma, adenoma, leiomyoma, rhabdomyoma, hemangioma,lymphangioma, osteoma, chondroma, lipoma and fibroma.

Diseases of Vasculitis—Vasculitic diseases are characterized byinflammation of the vessels. Infiltration of leukocytes leads todestruction of the vessel walls, and the complement pathway is believedto play a major role in initiating leukocyte migration as well as theresultant damage manifested at the site of inflammation (Vasculitis,Second Edition, Edited by Ball and Bridges, Oxford University Press, pp47-53, 2008). The compounds provided in the present invention can beused to treat leukoclastic vasculitis, Anti-neutrophil cytoplasmicantibody (ANCA) associated vasculitis, immune vasculitis Wegener'sgranulomatosis, microscopic polyangiitis, Churg-Strauss syndrome,Henoch-Schonlein purpura, polyateritis nodosa, Rapidly ProgressiveGlomerulonephritis (RPGN), cryoglobulinaemia, giant cell arteritis(GCA), Behcet's disease and Takayasu's arteritis (TAK).

HIV infection and AIDS—C5a receptor modulators provided herein may beused to inhibit HIV infection, delay AIDS progression or decrease theseverity of symptoms or HIV infection and AIDS.

Neurodegenerative disorders and related diseases—Within further aspects,C5a antagonists provided herein may be used to treat Alzheimer'sdisease, multiple sclerosis, and cognitive function decline associatedwith cardiopulmonary bypass surgery and related procedures.

In one embodiment of the invention, the compounds of the invention canbe used for the treatment of diseases selected from the group consistingof sepsis (and associated disorders), COPD, rheumatoid arthritis, lupusnephritis and multiple sclerosis.

Treatment methods provided herein include, in general, administration toa patient an effective amount of one or more compounds provided herein.Suitable patients include those patients suffering from or susceptibleto (i.e., prophylactic treatment) a disorder or disease identifiedherein. Typical patients for treatment as described herein includemammals, particularly primates, especially humans. Other suitablepatients include domesticated companion animals such as a dog, cat,horse, and the like, or a livestock animal such as cattle, pig, sheepand the like.

In general, treatment methods provided herein comprise administering toa patient an effective amount of a compound one or more compoundsprovided herein. In a preferred embodiment, the compound(s) of theinvention are preferably administered to a patient (e.g., a human)orally or topically. The effective amount may be an amount sufficient tomodulate C5a receptor activity and/or an amount sufficient to reduce oralleviate the symptoms presented by the patient. Preferably, the amountadministered is sufficient to yield a plasma concentration of thecompound (or its active metabolite, if the compound is a pro-drug) highenough to detectably inhibit white blood cell (e.g., neutrophil)chemotaxis in vitro. Treatment regimens may vary depending on thecompound used and the particular condition to be treated; for treatmentof most disorders, a frequency of administration of 4 times daily orless is preferred. In general, a dosage regimen of 2 times daily is morepreferred, with once a day dosing particularly preferred. It will beunderstood, however, that the specific dose level and treatment regimenfor any particular patient will depend upon a variety of factorsincluding the activity of the specific compound employed, the age, bodyweight, general health, sex, diet, time of administration, route ofadministration, rate of excretion, drug combination (i.e., other drugsbeing administered to the patient) and the severity of the particulardisease undergoing therapy, as well as the judgment of the prescribingmedical practitioner. In general, the use of the minimum dose sufficientto provide effective therapy is preferred. Patients may generally bemonitored for therapeutic effectiveness using medical or veterinarycriteria suitable for the condition being treated or prevented.

Dosage levels of the order of from about 0.1 mg to about 140 mg perkilogram of body weight per day are useful in the treatment orpreventions of conditions involving pathogenic C5a activity (about 0.5mg to about 7 g per human patient per day). The amount of activeingredient that may be combined with the carrier materials to produce asingle dosage form will vary depending upon the host treated and theparticular mode of administration. Dosage unit forms will generallycontain between from about 1 mg to about 500 mg of an active ingredient.For compounds administered orally, transdermally, intravaneously, orsubcutaneously, it is preferred that sufficient amount of the compoundbe administered to achieve a serum concentration of 5 ng(nanograms)/mL-10 μg (micrograms)/mL serum, more preferably sufficientcompound to achieve a serum concentration of 20 ng-1 μg/ml serum shouldbe administered, most preferably sufficient compound to achieve a serumconcentration of 50 ng/ml-200 ng/ml serum should be administered. Fordirect injection into the synovium (for the treatment of arthritis)sufficient compounds should be administered to achieve a localconcentration of approximately 1 micromolar.

Frequency of dosage may also vary depending on the compound used and theparticular disease treated. However, for treatment of most disorders, adosage regimen of 4 times daily, three times daily, or less ispreferred, with a dosage regimen of once daily or 2 times daily beingparticularly preferred. It will be understood, however, that thespecific dose level for any particular patient will depend upon avariety of factors including the activity of the specific compoundemployed, the age, body weight, general health, sex, diet, time ofadministration, route of administration, and rate of excretion, drugcombination (i.e., other drugs being administered to the patient), theseverity of the particular disease undergoing therapy, and otherfactors, including the judgment of the prescribing medical practitioner.

Combination Therapy

The presently disclosed compounds may be used in combination with one ormore additional therapeutic agents that are used in the treatment,prevention, suppression or amelioration of the diseases or conditionsfor which compounds and compositions of the present invention areuseful. Such one or more additional therapeutic agents may beadministered, by a route and in an amount commonly used therefor,contemporaneously or sequentially with a compound or composition of thepresent invention. When a compound or composition of the presentinvention is used contemporaneously with one or more other drugs, apharmaceutical composition containing such other drugs in addition tothe compound or composition of the present invention is preferred.Accordingly, the pharmaceutical compositions of the present inventioninclude those that also contain one or more other active ingredients ortherapeutic agents, in addition to a compound or composition of thepresent invention.

Examples of the the one or more additional therapeutic agents arecorticosteroids, steroids, immunosuppressants, Immunoglobulin Gagonists, Dipeptidyl peptidase IV inhibitors, Lymphocyte functionantigen-3 receptor antagonists, Interleukin-2 ligands, Interleukin-1beta ligand inhibitors, IL-2 receptor alpha subunit inhibitors, HGF genestimulators, IL-6 antagonists, IL-5 antagonists, Alpha 1 antitrypsinstimulators, Cannabinoid receptor antagonists, Histone deacetylaseinhibitors, AKT protein kinase inhibitors, CD20 inhibitors, Abl tyrosinekinase inhibitors, JAK tyrosine kinase inhibitors, TNF alpha ligandinhibitors, Hemoglobin modulators, TNF antagonists, proteasomeinhibitors, CD3 modulators, Hsp 70 family inhibitors, Immunoglobulinagonists, CD30 antagonists, tubulin antagonists, Sphingosine-1-phosphatereceptor-1 agonists, connective tissue growth factor ligand inhibitors,caspase inhibitors, adrenocorticotrophic hormone ligands, Btk tyrosinekinase inhibitors, Complement C1s subcomponent inhibitors,Erythropoietin receptor agonists, B-lymphocyte stimulator ligandinhibitors, Cyclin-dependent kinase-2 inhibitors, P-selectinglycoprotein ligand-1 stimulators, mTOR inhibitors, Elongation factor 2inhibitors, Cell adhesion molecule inhibitors, Factor XIII agonists,Calcineurin inhibitors, Immunoglobulin G1 agonists, Inosinemonophosphate dehydrogenase inhibitors, Complement C1s subcomponentinhibitors, Thymidine kinase modulators, Cytotoxic T-lymphocyteprotein-4 modulators, Angiotensin II receptor antagonists, AngiotensinII receptor modulators, TNF superfamily receptor 12A antagonists, CD52antagonists, Adenosine deaminase inhibitors, T-cell differentiationantigen CD6 inhibitors, FGF-7 ligands, dihydroorotate dehydrogenaseinhibitors, Syk tyrosine kinase inhibitors, Interferon type I receptorantagonists, Interferon alpha ligand inhibitors, Macrophage migrationinhibitory factor inhibitors, Integrin alpha-V/beta-6 antagonists,Cysteine protease stimulators, p38 MAP kinase inhibitors, TP53 geneinhibitors, Shiga like toxin I inhibitors, Fucosyltransferase 6stimulators, Interleukin 22 ligands, IRS1 gene inhibitors, Proteinkinase C stimulators, Protein kinase C alpha inhibitors, CD74antagonists, Immunoglobulin gamma Fc receptor IIB antagonists, T-cellantigen CD7 inhibitors, CD95 antagonists, N acetylmannosamine kinasestimulators, Cardiotrophin-1 ligands, Leukocyte elastase inhibitors,CD40 ligand receptor antagonists, CD40 ligand modulators, IL-17antagonists, TLR-2 antagonists, Mannan-binding lectin serine protease-2(MASP-2) inhibitors, Factor B inhibitors, Factor D inhibitors, C3aRmodulators, C5aR2 modulators, T cell receptor antagonists, PD-1inhibitors, PD-L1 inhibitors, TIGIT inhibitors, TIM-3 inhibitors, LAG-3inhibitors, VISTA inhibitors, STING agonists, IDO inhibitors, adenosinereceptor modulators, CD39 inhibitors, CD73 inhibitors, antagonists ofthe chemokine receptors, especially CXCR1, CXCR2, CXCR3, CXCR4, CXCR7,CCR1, CCR2, CCR3, CCR4, CCR5, CCR7, CCR7, CCR9, CX3CR1 and CXCR6, andcombinations thereof.

In some embodiments, the additional therapeutic agent used in thetherapeutic methods herein, is selected from the group consisting ofobinutuzumab, rituximab, ocrelizumab, cyclophosphamide, prednisone,hydrocortisone, hydrocortisone acetate, cortisone acetate, tixocortolpivalate, prednisolone, methylprednisolone, triamcinolone acetonide,triamcinolone alcohol, mometasone, amcinonide, budesonide, desonide,fluocinonide, fluocinolone acetonide, halcinonide, betamethasone,betamethasone sodium phosphate, dexamethasone, dexamethasone sodiumphosphate, fluocortolone, hydrocortisone-17-valerate, halometasone,alclometasone dipropionate, beclomethasone, betamethasone valerate,betamethasone dipropionate, prednicarbate, clobetasone-17-butyrate,clobetasol-17-propionate, fluocortolone caproate, fluocortolonepivalate, fluprednidene acetate, hydrocortisone-17-butyrate,hydrocortisone-17-aceponate, hydrocortisone-17-buteprate, ciclesonideand prednicarbate, GB-0998, immuglo, begelomab, alefacept, aldesleukin,gevokizumab, daclizumab, basiliximab, inolimomab, beperminogeneperplasmid, sirukumab, tocilizumab, clazakizumab, mepolizumab,fingolimod, panobinostat, triciribine, nilotinib, imatinib, tofacitinib,momelotinib, peficitinib, itacitinib, infliximab, PEG-bHb-CO,etanercept, ixazomib, bortezomib, muromonab, otelixizumab, gusperimus,brentuximab vedotin, Ponesimod, KRP-203, FG-3019, emricasan,corticotropin, ibrutinib, cinryze, conestat, methoxy polyethyleneglycol-epoetin beta, belimumab, blisibimod, atacicept, seliciclib,neihulizumab, everolimus, sirolimus, denileukin diftitox, LMB-2,natalizumab, catridecacog, ciclosporin, tacrolimus, voclosporin,voclosporin, canakinumab, mycophenolate, mizoribine, CE-1145, TK-DLI,abatacept, belatacept, olmesartan medoxomil, sparsentan, TXA-127,BIIB-023, alemtuzumab, pentostatin, itolizumab, palifermin, leflunomide,PRO-140, cenicriviroc, fostamatinib, anifrolumab, sifalimumab, BAX-069,BG-00011, losmapimod, QPI-1002, ShigamAbs, TZ-101, F-652, reparixin,ladarixin, PTX-9908, aganirsen, APH-703, sotrastaurin, sotrastaurin,milatuzumab, SM-101, T-Guard, APG-101, DEX-M74, cardiotrophin-1,tiprelestat, ASKP-1240, BMS-986004, HPH-116, KD-025, OPN-305, TOL-101,defibrotide, pomalidomide, Thymoglobulin, laquinimod, remestemcel-L,Equine antithymocyte immunoglobulin, Stempeucel, LIV-Gamma, Octagam 10%,t2c-001, 99mTc-sestamibi, Clairyg, Prosorba, pomalidomide, laquinimod,teplizumab, FCRx, solnatide, foralumab, ATIR-101, BPX-501, ACP-01,ALLO-ASC-DFU, irbesartan+propagermanium, ApoCell, cannabidiol, RGI-2001,saratin, anti-CD3 bivalent antibody-diphtheria toxin conjugate, NOX-100,LT-1951, OMS721, ALN-CC5, ACH-4471, AMY-101, Acthar gel, and CD4+CD25+regulatory T-cells, MEDI7814, P32, P59, pembrolizumab, nivolumab,atezolizumab, avelumab, durvalumab, CCX354, CCX721, CCX9588, CCX140,CCX872, CCX598, CCX6239, CCX587, CCX624, CCX282, CCX025, CCX507, CCX430,CCX765, CCX758, CCX771, CCX662, CCX650, and combinations thereof.

The disease or disorder being treated will determine which additionaltherapeutic agent or therapeutic agents are most appropriatelyadministered in combination with the compounds of the presentinvention—such determination can be made by a person of skill in theart.

The weight ratio of the compound of the present invention to the secondactive ingredient may be varied and will depend upon the effective doseof each ingredient. Generally, an effective dose of each will be used.Thus, for example, when a compound of the present invention is combinedwith an NSAID the weight ratio of the compound of the present inventionto the NSAID will generally range from about 1000:1 to about 1:1000,preferably about 200:1 to about 1:200. Combinations of a compound of thepresent invention and other active ingredients will generally also bewithin the aforementioned range, but in each case, an effective dose ofeach active ingredient should be used.

Non-Pharmaceutical Applications

In another aspect of the invention, the compounds of the invention canbe used in a variety of non-pharmaceutical in vitro and in vivoapplication. For example, the compounds of the invention may be labeledand used as probes for the detection and localization of C5a receptor(cell preparations or tissue sections samples). The compounds of theinvention may also be used as positive controls in assays for C5areceptor activity, i.e., as standards for determining the ability of acandidate agent to bind to C5a receptor, or as radiotracers for positronemission tomography (PET) imaging or for single photon emissioncomputerized tomography (SPECT). Such methods can be used tocharacterize C5a receptors in living subjects. For example, a C5areceptor modulator may be labeled using any of a variety of well knowntechniques (e.g., radiolabeled with a radionuclide such as tritium), andincubated with a sample for a suitable incubation time (e.g., determinedby first assaying a time course of binding). Following incubation,unbound compound is removed (e.g., by washing), and bound compounddetected using any method suitable for the label employed (e.g.,autoradiography or scintillation counting for radiolabeled compounds;spectroscopic methods may be used to detect luminescent groups andfluorescent groups). As a control, a matched sample containing labeledcompound and a greater (e.g., 10-fold greater) amount of unlabeledcompound may be processed in the same manner. A greater amount ofdetectable label remaining in the test sample than in the controlindicates the presence of C5a receptor in the sample. Detection assays,including receptor autoradiography (receptor mapping) of C5a receptor incultured cells or tissue samples may be performed as described by Kuharin sections 8.1.1 to 8.1.9 of Current Protocols in Pharmacology (1998)John Wiley & Sons, New York.

The compounds provided herein may also be used within a variety of wellknown cell separation methods. For example, modulators may be linked tothe interior surface of a tissue culture plate or other support, for useas affinity ligands for immobilizing and thereby isolating, C5areceptors (e.g., isolating receptor-expressing cells) in vitro. In onepreferred application, a modulator linked to a fluorescent marker, suchas fluorescein, is contacted with the cells, which are then analyzed (orisolated) by fluorescence activated cell sorting (FACS).

I. EXAMPLES

The following examples are offered to illustrate, but not to limit theclaimed invention.

Reagents and solvents used below can be obtained from commercial sourcessuch as Aldrich Chemical Co. (Milwaukee, Wis., USA). ¹H-NMR spectra wererecorded on a Varian Mercury 400 MHz NMR spectrometer. Significant peaksare provided relative to TMS and are tabulated in the order:multiplicity (s, singlet; d, doublet; t, triplet; q, quartet; m,multiplet) and number of protons. Mass spectrometry results are reportedas the ratio of mass over charge, followed by the relative abundance ofeach ion (in parenthesis). In the examples, a single m/e value isreported for the M+H (or, as noted, M−H) ion containing the most commonatomic isotopes. Isotope patterns correspond to the expected formula inall cases. Electrospray ionization (ESI) mass spectrometry analysis wasconducted on a Hewlett-Packard MSD electrospray mass spectrometer usingthe HP1100 HPLC for sample delivery. Normally the analyte was dissolvedin methanol at 0.1 mg/mL and 1 microliter was infused with the deliverysolvent into the mass spectrometer, which scanned from 100 to 1500daltons. All compounds could be analyzed in the positive ESI mode, usingacetonitrile/water with 1% formic acid as the delivery solvent. Thecompounds provided below could also be analyzed in the negative ESImode, using 2 mM NH₄OAc in acetonitrile/water as delivery system.

The following abbreviations are used in the Examples and throughout thedescription of the invention:

EtOH: Ethanol

EtONa: Sodium ethoxide

THF: Tetrahydrofuran

TLC: Thin layer chromatography

MeOH: Methanol

Compounds within the scope of this invention can be synthesized asdescribed below, using a variety of reactions known to the skilledartisan. One skilled in the art will also recognize that alternativemethods may be employed to synthesize the target compounds of thisinvention, and that the approaches described within the body of thisdocument are not exhaustive, but do provide broadly applicable andpractical routes to compounds of interest.

Certain molecules claimed in this patent can exist in differentenantiomeric and diastereomeric forms and all such variants of thesecompounds are claimed.

The detailed description of the experimental procedures used tosynthesize key compounds in this text lead to molecules that aredescribed by the physical data identifying them as well as by thestructural depictions associated with them.

Those skilled in the art will also recognize that during standard workup procedures in organic chemistry, acids and bases are frequently used.Salts of the parent compounds are sometimes produced, if they possessthe necessary intrinsic acidity or basicity, during the experimentalprocedures described within this patent.

Example 1 Synthesis of1-(4-(5-(2,4-bis(trifluoromethyl)benzyl)-2-(2,6-diethylphenyl)-6,6-dimethyl-2,4,5,6-tetrahydropyrrolo[3,4-c]pyrazol-3-yl)-2,5-difluorophenyl)urea

Caution: Diazonium formation could be potentially dangerous, pleasehandle with care and wear proper personal protective equipment!

Step a: To a 250 mL flask charged with 90 mL of concentratedhydrochloric acid under magnetic stirring was added 2,6-diethylaniline(10.0 g, 67.0 mmol). The resulting mixture was stirred for 30 min andcooled with an ice-salt bath until the internal temperature reached −5°C. A solution of sodium nitrite (5.5 g, 80.0 mmol) in water (60 mL) wasadded slowly to the above mixture while maintaining the internaltemperature below 5° C.

Separately, tin(II) chloride dihydrate (31.6 g, 140.0 mmol) was added toa 500 mL 3-neck round bottom flask charged with concentratedhydrochloric acid (60 mL) under mechanical stirring. The resultingsolution was then cooled with an ice bath.

The diazonium slurry was then filtered into the 500 mL flask containingthe cooled tin chloride solution with vigorous stirring. After 90 min,the reaction mixture was transferred to a 500 mL Erlenmeyer flask andthe flask was rinsed with water (20 mL) and chloroform (8 mL). Thecombined mixture was stirred overnight at room temperature. The entireliquid layer was decanted to give a wet solid. The recovered materialwas dried in vacuo for one day and then transferred to a 500 mL 3-neckround bottom flask equipped with an overhead mechanical stirrer andstirred with ether (180 mL). The resulting mixture was cooled in an icebath, and NaOH solution (10 N, 30 mL) was added slowly to the abovemixture while maintaining the inner temperature below 12° C. After theaddition, the mixture was allowed to stand for 2 h on ice. The etherlayer was decanted into a 500 mL flask and a stream of hydrogen chloridegas was bubbled into the ether solution while stirring. The resultingprecipitate was collected by filtration to afford(2,6-diethylphenyl)hydrazine hydrochloride. MS: (ES) m/z calculated forC₁₀H₁₇N₂ [M+H]⁺ 165.1, found 165.1.

Step b: Pyridine (4.0 mL, 49.5 mmol) was added to a mixture of(2,6-diethylphenyl)hydrazine hydrochloride (5.0 g, 24.9 mmol),tert-butyl 4-cyano-2,2-dimethyl-3-oxopyrrolidine-1-carboxylate (5.0 g,21.0 mmol) and EtOH (60 mL) in a 250 mL round bottom flask undermagnetic stirring. The resulting mixture was stirred at 70° C. for 24 h.The solvent was removed under reduced pressure, and the residue wasdiluted with EtOAc and washed with aqueous citric acid solution,saturated aqueous NaHCO₃ solution, brine, and dried over MgSO₄. Thesolvent was removed under reduced pressure and the residue wascrystallized from cyclohexane to give tert-butyl3-amino-2-(2,6-diethylphenyl)-6,6-dimethyl-2,6-dihydropyrrolo[3,4-c]pyrazole-5(4H)-carboxylate.MS: (ES) m/z calculated for C₂₂H₃₃N₄O₂ [M+H]⁺ 385.2, found 385.2.

Caution: Diazonium formation could be potentially dangerous, pleasehandle with care and wear proper personal protective equipment!

Tert-butyl nitrite (0.5 mL, 3.8 mmol) was added slowly at roomtemperature to a mixture of tert-butyl3-amino-2-(2,6-diethylphenyl)-6,6-dimethyl-2,6-dihydropyrrolo[3,4-c]pyrazole-5(4H)-carboxylate(1 g, 2.6 mmol), diiodomethane (1.5 mL, 18.6 mmol) and MeCN (15 mL) in a100 mL round bottom flask under magnetic stirring. The resulting mixturewas stirred at 45° C. for 3 h before it was diluted with toluene, washedwith saturated NH₄Cl solution/NH₄OH (3:1), brine, and dried over MgSO₄.The solvent was removed under reduced pressure and the residue waspurified by silica gel flash chromatography (2 to 25% EtOAc in hexanes)to give tert-butyl2-(2,6-diethylphenyl)-3-iodo-6,6-dimethyl-2,6-dihydropyrrolo[3,4-c]pyrazole-5(4H)-carboxylate.MS: (ES) m/z calculated for C₂₂H₃₁IN₃O₂ [M+H]⁺ 496.1, found 496.2.

Step c: A mixture of 4-bromo-2,5-difluoroaniline (1.5 g, 7.2 mmol),4,4,4′,4′,5,5,5′,5′-octamethyl-2,2′-bi(1,3,2-dioxaborolane) (2.2 g, 8.7mmol), KOAc (1.8 g, 18.3 mmol) and Pd(dppf)Cl₂ complex withdichloromethane (580.0 mg, 0.7 mmol) in dioxane (12 mL) was stirred at95° C. for 2 h under nitrogen. The mixture was then cooled to roomtemperature and filtered over Celite. The filtrate was collected,concentrated under reduced pressure, and purified by silica gel flashchromatography (0 to 50% EtOAc in hexanes) to give2,5-difluoro-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)aniline. MS:(ES) m/z calculated for C₁₂H₁₇BF₂NO₂ [M+H]⁺ 256.1, found 256.2.

To a suspension of tert-butyl2-(2,6-diethylphenyl)-3-iodo-6,6-dimethyl-2,6-dihydropyrrolo[3,4-c]pyrazole-5(4H)-carboxylate(0.7 g, 1.4 mmol),2,5-difluoro-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)aniline (0.7g, 2.7 mmol), K₂CO₃ (1.3 g, 7.2 mmol) in dioxane (10 mL) and water (2mL) was added Pd(dppf)Cl₂ complex with dichloromethane (300.0 mg, 0.37mmol). The reaction mixture was degassed (N₂) for 2 min and stirredunder N₂ at 100° C. for 2 h. The reaction mixture was diluted withEtOAc, filtered through Celite, washed with brine, dried over MgSO₄, andfiltered. The solvent was removed under reduced pressure and the residuewas purified by silica gel flash chromatography (2 to 10% EtOAc inhexanes) to give tert-butyl3-(4-amino-2,5-difluorophenyl)-2-(2,6-diethylphenyl)-6,6-dimethyl-2,6-dihydropyrrolo[3,4-c]pyrazole-5(4H)-carboxylate.MS: (ES) m/z calculated for C₂₈H₃₅F₂N₄O₂ [M+H]⁺ 497.3, found 497.5.

Step d: A mixture of tert-butyl3-(4-amino-2,5-difluorophenyl)-2-(2,6-diethylphenyl)-6,6-dimethyl-2,6-dihydropyrrolo[3,4-c]pyrazole-5(4H)-carboxylate(0.5 g, 1.0 mmol) and benzoyl isocyanate (0.5 g, 3.4 mmol) in THF (10mL) was stirred for 3 h at room temperature. The mixture wasconcentrated under reduced pressure to obtain tert-butyl3-(4-(3-benzoylureido)-2,5-difluorophenyl)-2-(2,6-diethylphenyl)-6,6-dimethyl-2,6-dihydropyrrolo[3,4-c]pyrazole-5(4H)-carboxylate.

A mixture of tert-butyl3-(4-(3-benzoylureido)-2,5-difluorophenyl)-2-(2,6-diethylphenyl)-6,6-dimethyl-2,6-dihydropyrrolo[3,4-c]pyrazole-5(4H)-carboxylate(˜1.0 mmol, from above) and K₂CO₃ (1.3 g, 7.2 mmol) in MeOH (15 mL) wasstirred for 2 h at room temperature followed by 20 min at 50° C. Themixture was extracted with EtOAc. The organic layer was separated, driedover MgSO₄, concentrated under reduced pressure and purified by silicagel flash chromatography (10 to 50% EtOAc in hexanes) to affordtert-butyl2-(2,6-diethylphenyl)-3-(2,5-difluoro-4-ureidophenyl)-6,6-dimethyl-2,6-dihydropyrrolo[3,4-c]pyrazole-5(4H)-carboxylate.MS: (ES) m/z calculated for C₂₉H₃₆F₂N₅O₃ [M+H]⁺ 540.3, found 540.3.

Step e: The above tert-butyl2-(2,6-diethylphenyl)-3-iodo-6,6-dimethyl-2,6-dihydropyrrolo[3,4-c]pyrazole-5(4H)-carboxylatewas dissolved in dichloromethane (10 mL) and charged with HCl in dioxane(4 N, 5 mL). The resulting mixture was stirred at room temperature for12 h. Upon completion, the solvent was evaporated in vacuo to give1-(4-(2-(2,6-diethylphenyl)-6,6-dimethyl-2,4,5,6-tetrahydropyrrolo[3,4-c]pyrazol-3-yl)-2,5-difluorophenyl)ureahydrochloride. MS: (ES) m/z calculated for C₂₄H₂₈F₂N₅O [M+H]⁺ 440.2,found 440.3.

Step f: N,N-diisopropylethylamine (0.2 mL, 1.2 mmol) was added to asuspension1-(4-(2-(2,6-diethylphenyl)-6,6-dimethyl-2,4,5,6-tetrahydropyrrolo[3,4-c]pyrazol-3-yl)-2,5-difluorophenyl)ureahydrochloride (0.1 g, 0.2 mmol), and2,4-bis(trifluoromethyl)benzaldehyde (0.2 g, 0.8 mmol) in1,2-dichloroethane (10 mL) under magnetic stirring. After stirring atroom temperature for 10 min, NaBH(OAc)₃ (0.3 g, 1.4 mmol) was added inportions. The resulting mixture was stirred at 45° C. for 2 h. Aftercooling to room temperature, the reaction mixture was diluted withEtOAc, washed with aqueous NaHCO₃ solution, brine and dried over MgSO₄.The solvent was removed under reduced pressure and the residue waspurified by preparative TLC (50% EtOAc in hexanes) followed by HPLC(MeCN/H₂O, with 1% TFA) to give1-(4-(5-(2,4-bis(trifluoromethyl)benzyl)-2-(2,6-diethylphenyl)-6,6-dimethyl-2,4,5,6-tetrahydropyrrolo[3,4-c]pyrazol-3-yl)-2,5-difluorophenyl)urea.¹H NMR (400 MHz, CDCl₃) δ 8.18 (d, J=8.3 Hz, 1H), 7.88-7.98 (m, 2H),7.75-7.83 (m, 1H), 7.31 (t, J=7.7 Hz, 1H), 7.14 (d, J=7.7 Hz, 2H),6.79-6.85 (br, 1H), 6.40 (dd, J=6.5, 12.1 Hz, 1H), 4.79 (s, 2H), 4.13(s, 2H), 3.74 (s, 2H), 2.20-2.34 (m, 4H), 1.51 (s, 6H), 1.06 (t, J=7.6Hz, 6H). MS: (ES) m/z calculated for C₃₃H₃₂F₈N₅O [M+H]⁺ 666.2, found666.2.

Example 2 Synthesis of1-(4-(2-(2,6-diethylphenyl)-5-isobutyryl-6,6-dimethyl-2,4,5,6-tetrahydropyrrolo[3,4-c]pyrazol-3-yl)-2,5-difluorophenyl)urea

A mixture of4-(2-(2,6-diethylphenyl)-6,6-dimethyl-2,4,5,6-tetrahydropyrrolo[3,4-c]pyrazol-3-yl)-2,5-difluoroanilinehydrochloride (30.0 mg, 0.06 mmol), isobutyric acid (44.0 mg, 0.5 mmol),HATU (190.0 mg, 0.5 mmol) and NEt₃ (0.10 mL, 0.7 mmol) in DMF (1.5 mL)was stirred at 50° C. for 30 min. The mixture was then cooled to roomtemperature, quenched with saturated aqueous NaHCO₃ solution andextracted with EtOAc. The organic layer was separated, washed withbrine, dried over Na₂SO₄ and filtered. The solvent was concentratedunder reduced pressure and the residue was purified by silica gel flashchromatography (0 to 100% EtOAc in hexanes) to afford1-(4-(2-(2,6-diethylphenyl)-5-isobutyryl-6,6-dimethyl-2,4,5,6-tetrahydropyrrolo[3,4-c]pyrazol-3-yl)-2,5-difluorophenyl)urea.¹H NMR (400 MHz, CD₃OD) δ 8.06 (m, 1H), 7.44 (dd, J=7.8, 7.8 Hz, 1H),7.26 (d, J=7.6 Hz, 2H), 6.50 (m, 1H), 4.83 (s, 2H), 3.30 (m, 3H), 2.84(septet, J=6.6 Hz, 1H), 2.27 (m, 4H), 1.82 (s, 6H), 1.15 (d, J=6.4 Hz,6H), 1.06 (t, J=7.6 Hz, 6H). MS: (ES) m/z calculated for C₂₈H₃₄F₂N₅O₂[M+H]⁺ 510.3, found 510.6.

Example 3 Synthesis of1-(4-(5-(2,4-bis(trifluoromethyl)benzyl)-2-(2,6-diethylphenyl)-6,6-dimethyl-2,4,5,6-tetrahydropyrrolo[3,4-c]pyrazol-3-yl)-5-fluoro-2-methylphenyl)urea

Step a: Tert-butyl2-(2,6-diethylphenyl)-3-iodo-6,6-dimethyl-2,6-dihydropyrrolo[3,4-c]pyrazole-5(4H)-carboxylate(1.0 g, 2.0 mmol) was dissolved in dichloromethane (10 mL) and chargedwith HCl in dioxane (4 N, 5 mL). The resulting mixture was stirred atroom temperature for 12 h. After the reaction was complete, the solventwas evaporated in vacuo to give2-(2,6-diethylphenyl)-3-iodo-6,6-dimethyl-2,4,5,6-tetrahydropyrrolo[3,4-c]pyrazolehydrochloride. MS: (ES) m/z calculated for C₁₇H₂₃IN₃ [M+H]⁺ 396.1, found396.2.

N,N-diisopropylethylamine (0.3 mL, 1.7 mmol) was added to a suspensionof2-(2,6-diethylphenyl)-3-iodo-6,6-dimethyl-2,4,5,6-tetrahydropyrrolo[3,4-c]pyrazolehydrochloride (680.0 mg, 1.6 mmol) and2,4-bis(trifluoromethyl)benzaldehyde (0.8 g, 3.3 mmol) in1,2-dichloroethane (10 mL) under magnetic stirring. After stirring atroom temperature for 10 min, NaBH(OAc)₃ (0.8 g, 3.8 mmol) was added inportions. The resulting mixture was stirred at 45° C. for 2 h. Aftercooling to room temperature, the reaction mixture was diluted withEtOAc, washed with aqueous NaHCO₃ solution, brine and dried over MgSO₄.The solvent was removed under reduced pressure and the residue waspurified by silica gel flash chromatography (2 to 25% EtOAc in hexanes)to give5-(2,4-bis(trifluoromethyl)benzyl)-2-(2,6-diethylphenyl)-3-iodo-6,6-dimethyl-2,4,5,6-tetrahydropyrrolo[3,4-c]pyrazole.MS: (ES) m/z calculated for C₂₆H₂₇F₆IN₃ [M+H]⁺ 622.1, found 622.1.

Step b: A mixture of 4-bromo-5-fluoro-2-methylaniline (1.5 g, 7.4 mmol),4,4,4′,4′,5,5,5′,5′-octamethyl-2,2′-bi(1,3,2-dioxaborolane) (2.2 g, 8.7mmol), KOAc (1.8 g, 18.3 mmol) and Pd(dppf)Cl₂ complex withdichloromethane (580.0 mg, 0.7 mmol) in dioxane (12 mL) was stirred at95° C. for 2 h under nitrogen. The mixture was then cooled to roomtemperature and filtered over Celite. The filtrate was collected,concentrated under reduced pressure, and purified by silica gel flashchromatography (0 to 50% EtOAc in hexanes) to give5-fluoro-2-methyl-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)aniline.MS: (ES) m/z calculated for C₁₃H₂₀BFNO₂ [M+H]⁺ 252.2, found 252.2.

To a suspension of5-(2,4-bis(trifluoromethyl)benzyl)-2-(2,6-diethylphenyl)-3-iodo-6,6-dimethyl-2,4,5,6-tetrahydropyrrolo[3,4-c]pyrazole(75.0 mg, 0.12 mmol),5-fluoro-2-methyl-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)aniline(100.0 mg, 0.4 mmol), and K₂CO₃ (445.0 mg, 1.8 mmol) in dioxane (6 mL)and water (1 mL) was added Pd(dppf)Cl₂ complex with dichloromethane (50mg, 0.06 mmol). The reaction mixture was degassed (N₂) for 2 min andstirred under N₂ at 100° C. for 2.5 h. The reaction mixture was dilutedwith EtOAc, washed with aqueous NaHCO₃ solution and dried over Na₂SO₄.The solvent was removed under reduced pressure and the residue waspurified by silica gel flash chromatography (3 to 30% EtOAc in hexanes)to give4-(5-(2,4-bis(trifluoromethyl)benzyl)-2-(2,6-diethylphenyl)-6,6-dimethyl-2,4,5,6-tetrahydropyrrolo[3,4-c]pyrazol-3-yl)-5-fluoro-2-methylaniline.MS: (ES) m/z calculated for C₃₃H₃₄F₇N₄ [M+H]⁺ 619.3, found 619.3.

Step c: A mixture of the above4-(5-(2,4-bis(trifluoromethyl)benzyl)-2-(2,6-diethylphenyl)-6,6-dimethyl-2,4,5,6-tetrahydropyrrlo[3,4-c]pyrazol-3-yl)-5-fluoro-2-methylaniline(˜0.1 mmol) and benzoyl isocyanate (100.0 mg, 0.7 mmol) in THF (10 mL)was stirred for 3 h at room temperature. The mixture was concentratedunder reduced pressure to obtainN-((4-(5-(2,4-bis(trifluoromethyl)benzyl)-2-(2,6-diethylphenyl)-6,6-dimethyl-2,4,5,6-tetrahydropyrrolo[3,4-c]pyrazol-3-yl)-5-fluoro-2-methylphenyl)carbamoyl)benzamide.

A mixture ofN-((4-(5-(2,4-bis(trifluoromethyl)benzyl)-2-(2,6-diethylphenyl)-6,6-dimethyl-2,4,5,6-tetrahydropyrrolo[3,4-c]pyrazol-3-yl)-5-fluoro-2-methylphenyl)carbamoyl)benzamide(˜0.1 mmol, from above) and K₂CO₃ (138.0 mg, 1.0 mmol) in MeOH (6 mL)was stirred for 2 h at room temperature followed by 20 min at 50° C. Themixture was extracted with EtOAc. The organic layer was separated, driedover MgSO₄, concentrated under reduced pressure and purified bypreparative TLC (50% EtOAc in hexanes) followed by HPLC (MeCN/H₂O, with1% TFA) to give1-(4-(5-(2,4-bis(trifluoromethyl)benzyl)-2-(2,6-diethylphenyl)-6,6-dimethyl-2,4,5,6-tetrahydropyrro[3,4-c]pyrazol-3-yl)-5-fluoro-2-methylphenyl)urea.¹H NMR (400 MHz, CD₃OD): δ 8.24 (d, J=8.3 Hz, 1H), 7.93-7.98 (m, 2H),7.66 (d, J=13.4 Hz, 1H), 7.39 (dd, J=7.2, 8.1 Hz, 1H), 7.22 (d, J=7.7Hz, 2H), 6.56 (d, J=8.1 Hz, 1H), 4.87 (br, 3H), 4.20 (s, 2H), 3.73 (s,2H), 2.20-2.34 (m, 4H), 1.90 (s, 3H), 1.52 (s, 6H), 1.07 (t, J=7.6 Hz,6H). MS: (ES) m/z calculated for C₃₄H₃₅F₇N₅O [M+H]⁺ 662.3, found 662.5.

Example 4 Synthesis of1-(4-(5-(2,4-bis(trifluoromethyl)benzyl)-2-(2,6-diethylphenyl)-6,6-dimethyl-2,4,5,6-tetrahydropyrrolo[3,4-c]pyrazol-3-yl)-2-chloro-5-fluorophenyl)urea

Step a: To a suspension of 4-bromo-2-chloro-5-fluoroaniline (1.05 g, 4.7mmol), 4,4,4′,4′,5,5,5′,5′-octamethyl-2,2′-bi(1,3,2-dioxaborolane) (1.3g, 5.1 mmol) and KOAc (1.2 g, 11.7 mmol) in dioxane (15 mL) was addedPd(dppf)Cl₂ complex with dichloromethane (417.0 mg, 0.5 mmol). Themixture was degassed (N₂) for 2 min and stirred at 95° C. for 2.5 h. Themixture was cooled to room temperature, diluted with EtOAc and filteredthrough Celite. The filtrate was collected and concentrated underreduced pressure. The residue was purified by silica gel flashchromatography (0 to 40% EtOAc in hexanes) to give2-chloro-5-fluoro-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)aniline.MS: (ES) m/z calculated for C₁₂H₁₇BClFNO₂ [M+H]⁺ 272.1, found 272.1.

A mixture of5-(2,4-bis(trifluoromethyl)benzyl)-2-(2,6-diethylphenyl)-3-iodo-6,6-dimethyl-2,4,5,6-tetrahydropyrrolo[3,4-c]pyrazole(50.0 mg, 0.08 mmol),2-chloro-5-fluoro-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)aniline(93.0 mg, 0.3 mmol), K₂CO₃ (170.0 mg, 1.2 mmol) and Pd(dppf)Cl₂ complexwith dichloromethane (60.0 mg, 0.07 mmol) in dioxane (3 mL) and water(0.5 mL) was stirred at 100° C. for 1 h under N₂. It was then cooled toroom temperature, quenched with saturated NaHCO₃ solution and extractedwith EtOAc. The organic layer was separated, washed with brine, driedover Na₂SO₄, filtered, and concentrated under reduced pressure. Theresidue was purified by silica gel flash chromatography (0 to 50% EtOAcin hexanes) to afford4-(5-(2,4-bis(trifluoromethyl)benzyl)-2-(2,6-diethylphenyl)-6,6-dimethyl-2,4,5,6-tetrahydropyrrolo[3,4-c]pyrazol-3-yl)-2-chloro-5-fluoroaniline.MS: (ES) m/z calculated for C₃₂H₃₁ClF₈N₄ [M+H]⁺ 639.2, found 639.2.

Step b: A mixture of4-(5-(2,4-bis(trifluoromethyl)benzyl)-2-(2,6-diethylphenyl)-6,6-dimethyl-2,4,5,6-tetrahydropyrrolo[3,4-c]pyrazol-3-yl)-2-chloro-5-fluoroaniline(35.0 mg, 0.06 mmol) and benzoyl isocyanate (40.0 mg, 0.27 mmol) in THF(3 mL) was stirred at room temperature for 2 h. It was concentratedunder reduced pressure to affordN-((4-(5-(2,4-bis(trifluoromethyl)benzyl)-2-(2,6-diethylphenyl)-6,6-dimethyl-2,4,5,6-tetrahydropyrrolo[3,4-c]pyrazol-3-yl)-2-chloro-5-fluorophenyl)carbamoyl)benzamide.

A mixture of the aboveN-((4-(5-(2,4-bis(trifluoromethyl)benzyl)-2-(2,6-diethylphenyl)-6,6-dimethyl-2,4,5,6-tetrahydropyrrolo[3,4-c]pyrazol-3-yl)-2-chloro-5-fluorophenyl)carbamoyl)benzamide(˜0.06 mmol) and K₂CO₃ (200.0 mg, 1.2 mmol) in MeOH (5 mL) was stirredat 30° C. for 1 h. The mixture was then cooled to room temperature,quenched with saturated NaHCO₃ solution and extracted with EtOAc. Theorganic layer was separated, washed with brine, dried over Na₂SO₄,filtered, and concentrated under reduced pressure. The residue waspurified by silica gel flash chromatography (0 to 90% EtOAc in hexanes)followed by HPLC (MeCN/H₂O, with 1% TFA). The pure HPLC fraction wasbasified with saturated NaHCO₃ solution and extracted with EtOAc. Theorganic layer was separated, dried over Na₂SO₄ and concentrated underreduced pressure. The obtained material was treated with 1 M HCl inether (0.2 mL) and evaporated to dryness to afford1-(4-(5-(2,4-bis(trifluoromethyl)benzyl)-2-(2,6-diethylphenyl)-6,6-dimethyl-2,4,5,6-tetrahydropyrrolo[3,4-c]pyrazol-3-yl)-2-chloro-5-fluorophenyl)ureaas the HCl salt. ¹H NMR (HCl salt) (400 MHz, CD₃OD) δ 8.15 (m, 5H), 7.47(dd, J=7.8, 7.8 Hz, 1H), 7.28 (d, J=8.0, 2H), 6.70 (d, J=7.6 Hz, 1H),4.30-5.00 (m, 5H), 2.24 (m, 4H), 1.92 (br s, 6H), 1.28 (m, 2H), 1.05 (m,6H). MS (free form): (ES) m/z calculated for C₃₃H₃₂ClF₇N₅O [M+H]⁺ 682.2,found 682.2.

Example 5 Synthesis of1-(4-(2-(2,6-diethylphenyl)-6,6-dimethyl-5-(3,3,3-trifluoro-2,2-dimethylpropyl)-2,4,5,6-tetrahydropyrrolo[3,4-c]pyrazol-3-yl)-2,5-difluorophenyl)urea

Step a: A mixture of 3,3,3-trifluoro-2,2-dimethylpropanoic acid (468.0mg, 3.0 mmol), oxalyl chloride (0.25 mL, 3.0 mmol) and DMF (2 drops) inDCM (10 mL) was stirred at room temperature for 0.5 h to obtain3,3,3-trifluoro-2,2-dimethylpropanoyl chloride. The crude product wasdirectly used for the next step without further processing.

Step b: A mixture of 3,3,3-trifluoro-2,2-dimethylpropanoyl chloride(˜1.5 mmol) (obtained from step a above),2-(2,6-diethylphenyl)-3-iodo-6,6-dimethyl-2,4,5,6-tetrahydropyrrolo[3,4-c]pyrazolehydrochloride (253.0 mg, 0.58 mmol) and NEt₃ (0.41 mL, 2.9 mmol) and inDCM (5 mL) was stirred at room temperature for 1 h. It was quenched withsaturated aqueous NaHCO₃ solution and extracted with EtOAc. The organiclayer was separated, washed with brine, dried over Na₂SO₄ and filtered.The solvent was concentrated under reduced pressure and the residue waspurified by silica gel flash chromatography (0 to 60% EtOAc in hexanes)to afford1-(2-(2,6-diethylphenyl)-3-iodo-6,6-dimethyl-2,6-dihydropyrrolo[3,4-c]pyrazol-5(4H)-yl)-3,3,3-trifluoro-2,2-dimethylpropan-1-one,which was used directly for next step. MS: (ES) m/z calculated forC₂₆H₂₈F₃IN₃O [M+H]⁺ 534.1, found 534.1.

Step c: A mixture of1-(2-(2,6-diethylphenyl)-3-iodo-6,6-dimethyl-2,6-dihydropyrrolo[3,4-c]pyrazol-5(4H)-yl)-3,3,3-trifluoro-2,2-dimethylpropan-1-one(250.0 mg, ˜0.23 mmol, purity ˜50%),2,5-difluoro-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)aniline(320.0 mg, 1.25 mmol), K₂CO₃ (0.6 g, 4.3 mmol) and Pd(dppf)Cl₂ complexwith dichloromethane (120.0 mg, 0.15 mmol) in p-dioxane (12 mL) andwater (2 mL) was stirred at 100° C. for 1 h under N₂. It was then cooledto room temperature, quenched with saturated NaHCO₃ solution andextracted with EtOAc. The organic layer was separated, washed withbrine, dried over Na₂SO₄ and concentrated under reduced pressure. Theresidue was purified by silica gel flash chromatography (0 to 80% EtOAcin hexanes) to afford1-(3-(4-amino-2,5-difluorophenyl)-2-(2,6-diethylphenyl)-6,6-dimethyl-2,6-dihydropyrrolo[3,4-c]pyrazol-5(4H)-yl)-3,3,3-trifluoro-2,2-dimethylpropan-1-one. MS: (ES) m/zcalculated for C₂₈H₃₂F₅N₄O [M+H]⁺ 535.2, found 535.2.

Step d: To a solution of1-(3-(4-amino-2,5-difluorophenyl)-2-(2,6-diethylphenyl)-6,6-dimethyl-2,6-dihydropyrrolo[3,4-c]pyrazol-5(4H)-yl)-3,3,3-trifluoro-2,2-dimethylpropan-1-one(140.0 mg, 0.26 mmol) in THF (4 mL) was added 1 M DIBAL-H solution indichloromethane (2.5 mL, 2.5 mmol). The mixture was stirred at rt for 10min, quenched with water, basified with saturated NaHCO₃ solution andextracted with EtOAc. The organic layer was separated, washed withbrine, dried over Na₂SO₄ and concentrated under reduced pressure. Theresidue was purified by silica gel flash chromatography (0 to 60% EtOAcin hexanes) to afford4-(2-(2,6-diethylphenyl)-6,6-dimethyl-5-(3,3,3-trifluoro-2,2-dimethylpropyl)-2,4,5,6-tetrahydropyrrolo[3,4-c]pyrazol-3-yl)-2,5-difluoroaniline.MS: (ES) m/z calculated for C₂₈H₃₄F₅N₄ [M+H]⁺ 521.3, found 521.3.

Step e: A mixture of4-(2-(2,6-diethylphenyl)-6,6-dimethyl-5-(3,3,3-trifluoro-2,2-dimethylpropyl)-2,4,5,6-tetrahydropyrrolo[3,4-c]pyrazol-3-yl)-2,5-difluoroaniline(0.036 g, 0.069 mmol) and benzoyl isocyanate (30.0 mg, 0.2 mmol) in THF(3.5 mL) was stirred at room temperature for 0.5 h. It was concentratedunder reduced pressure to affordN-((4-(2-(2,6-diethylphenyl)-6,6-dimethyl-5-(3,3,3-trifluoro-2,2-dimethylpropyl)-2,4,5,6-tetrahydropyrrolo[3,4-c]pyrazol-3-yl)-2,5-difluorophenyl)carbamoyl)benzamide.

A mixture ofN-((4-(2-(2,6-diethylphenyl)-6,6-dimethyl-5-(3,3,3-trifluoro-2,2-dimethylpropyl)-2,4,5,6-tetrahydropyrrolo[3,4-c]pyrazol-3-yl)-2,5-difluorophenyl)carbamoyl)benzamide(˜0.07 mmol) and K₂CO₃ (200.0 mg, 1.2 mmol) in MeOH (4 mL) was stirredat room temperature for 1.5 h. It was then poured into saturated aqueousNaHCO₃ solution and extracted with EtOAc. The organic layer wasseparated, washed with brine, dried over Na₂SO₄ and concentrated underreduced pressure. The residue was purified by silica gel flashchromatography (0 to 90% EtOAc in hexanes) followed by HPLC (MeCN/H₂O,with 1% TFA) to afford1-(4-(2-(2,6-diethylphenyl)-6,6-dimethyl-5-(3,3,3-trifluoro-2,2-dimethylpropyl)-2,4,5,6-tetrahydropyrrolo[3,4-c]pyrazol-3-yl)-2,5-difluorophenyl)urea.¹H NMR (400 MHz, CDCl₃) δ 9.80 (d, J=2.4 Hz, 1H), 8.08 (m, 1H), 7.30(dd, J=7.8 Hz, 1H), 7.22 (br s, 1H), 7.10 (d, J=7.2 Hz, 2H), 6.64 (m,1H), 4.98 (s, 2H), 2.33 (s, 2H), 2.20 (m, 5H), 1.55 (s, 6H), 1.09 (m,12H). MS: (ES) m/z calculated for C₂₉H₃₅F₅N₅O [M+H]⁺ 564.2, found 564.2.

Example 6 Synthesis of4-(2-(2,6-dimethylphenyl)-5-(4-fluoro-5-isopropyl-2-methylphenyl)-2,4,5,6-tetrahydropyrrolo[3,4-c]pyrazol-3-yl)benzamide

Caution: Diazonium formation could be potentially dangerous, pleasehandle with care and wear proper personal protective equipment!

Step a: To tert-butyl 3-cyano-4-oxo-pyrrolidine-1-carboxylate (19.4 g,92.3 mmol) and (2,6-dimethylphenyl)hydrazine hydrochloride (16.0 g, 92.7mmol) were added EtOH (160 mL) and AcOH (40 mL). The resultingsuspension was stirred at 50° C. overnight. Upon completion, thereaction mixture was quenched with 1 N NaOH aqueous solution andextracted with EtOAc, dried over MgSO₄, filtered, and concentrated invacuo. The crude product was then purified by silica gel flashchromatography (50% EtOAc in hexanes) to obtain tert-butyl3-amino-2-(2,6-dimethylphenyl)-4,6-dihydropyrrolo[3,4-c]pyrazole-5-carboxylate.MS: (ES) m/z calculated for C₁₈H₂₅N₄O₂ [M+H]⁺ 329.2, found 329.2.

Step b: Isoamyl nitrite (11.7 mL, 87.5 mmol) was added slowly at roomtemperature to a mixture of tert-butyl3-amino-2-(2,6-dimethylphenyl)-4,6-dihydropyrrolo[3,4-c]pyrazole-5-carboxylate(14.4 g, 43.8 mmol), diiodomethane (14 mL, 175.0 mmol) and MeCN (180mL). The resulting reaction mixture was stirred at room temperature for2 h. The reaction mixture was purified by silica gel flashchromatography (30% EtOAc in hexanes) to obtain tert-butyl2-(2,6-dimethylphenyl)-3-iodo-4,6-dihydropyrrolo[3,4-c]pyrazole-5-carboxylate.MS: (ES) m/z calculated for C18H₂₃IN₃O₂ [M+H]⁺ 440.1, found 440.2.

Step c: To a suspension of tert-butyl2-(2,6-dimethylphenyl)-3-iodo-4,6-dihydropyrrolo[3,4-c]pyrazole-5-carboxylate(1.0 g, 2.3 mmol), (4-cyanophenyl)boronic acid (0.6 g, 4.1 mmol),Pd(dppf)Cl₂ complex with dichloromethane (150 mg, 0.18 mmol) and K₂CO₃(1.2 g, 8.7 mmol) in dioxane (10 mL) and water (2 mL) was stirred at 98°C. for 2 h under N₂. The mixture was cooled to room temperature,quenched with water, and extracted with EtOAc. The organic layer wasseparated, washed with brine, dried over MgSO₄, and filtered. Thesolvent was concentrated under reduced pressure and the residue waspurified by silica gel flash chromatography (2 to 25% EtOAc in hexanes)to yield tert-butyl3-(4-cyanophenyl)-2-(2,6-dimethylphenyl)-2,6-dihydropyrrolo[3,4-c]pyrazole-5(4H)-carboxylate.MS: (ES) m/z calculated for C₂₅H₂₇N₄O₂ [M+H]⁺ 415.2, found 415.2.

Step d: A mixture of tert-butyl3-(4-cyanophenyl)-2-(2,6-dimethylphenyl)-2,6-dihydropyrrolo[3,4-c]pyrazole-5(4H)-carboxylate(800.0 mg, 1.9 mmol) and 4 N HCl in dioxane (5.0 mL, 20.0 mmol) indichloromethane (5 mL) was stirred at room temperature for 1 h. Themixture was basified with saturated aqueous NaHCO₃ solution andextracted with EtOAc. The organic layer was separated, washed withbrine, dried over MgSO₄, and filtered. The solvent was concentratedunder reduced pressure and the residue was purified by silica gel flashchromatography (0 to 25% MeOH in dichloromethane) to yield4-(2-(2,6-dimethylphenyl)-2,4,5,6-tetrahydropyrrolo[3,4-c]pyrazol-3-yl)benzonitrile.MS: (ES) m/z calculated for C₂₀H₁₉N₄ [M+H]⁺ 315.2, found 315.2.

Step e: A mixture of 1-bromo-2-fluoro-4-methyl-5-nitrobenzene (2.0 g,8.5 mmol), 4,4,4′,4′,5,5,5′,5′-octamethyl-2,2′-bi(1,3,2-dioxaborolane)(2.0 g, 11.9 mmol, Pd(PPh₃)₄ (260.0 mg, 0.2 mmol), and K₂CO₃ (2.4 g,17.4 mmol) in dioxane (10 mL) and water (2 mL) was stirred at 98° C. for2 h under N₂. The mixture was cooled to room temperature, quenched withwater, and extracted with EtOAc. The organic layer was separated, washedwith brine, dried over MgSO₄, and filtered. The solvent was concentratedunder reduced pressure and the residue was purified by silica gel flashchromatography (2 to 10% EtOAc in hexanes) to yield1-fluoro-5-methyl-4-nitro-2-(prop-1-en-2-yl)benzene, which was directlyused for the next step.

A pressure vessel containing1-fluoro-5-methyl-4-nitro-2-(prop-1-en-2-yl)benzene (2.0 g, 10.2 mmol),10% Pd/C (2.0 g, 50% wet), EtOH (50 mL) and dichloromethane (50 mL) wasagitated under a hydrogen atmosphere at 45 psi for 3 h. The mixture wasfiltered through Celite. The filtrate was collected and concentratedunder reduced pressure. The residue was purified by silica gel flashchromatography (2 to 15% EtOAc in hexanes) to yield4-fluoro-5-isopropyl-2-methylaniline. C₁₀H₁₅FN [M+H]⁺ 168.1, found168.2.

Caution: Diazonium formation could be potentially dangerous, pleasehandle with care and wear proper personal protective equipment!

To a mixture of NaNO₂ (1.2 g, 17.7 mmol), CuBr (1.5 g, 10.5 mmol) inDMSO (20 mL) at 0° C. was added 4-fluoro-5-isopropyl-2-methylaniline(0.6 g, 3.6 mmol) followed by dropwise addition of aqueous 48% HBrsolution (4 mL). The obtained mixture was then stirred and allowed towarm up to room temperature over 1 h. The mixture was then poured intoice and extracted with EtOAc. The organic layer was separated, washedwith brine, dried over MgSO₄, and filtered. The solvent was concentratedunder reduced pressure and the residue was purified by silica gel flashchromatography (2 to 25% EtOAc in hexanes) to yield1-bromo-4-fluoro-5-isopropyl-2-methylbenzene, which was directly usedfor the next step. ¹H NMR (400 MHz, CDCl₃) δ 7.36 (d, J=8.0 Hz, 1H),6.89 (dt, J=0.6, 10.8 Hz, 1), 3.15 (hept, J=6.9 Hz, 1H), 2.33 (s, 3H),1.23 (d, J=6.9 Hz, 6H).

A mixture of4-(2-(2,6-dimethylphenyl)-2,4,5,6-tetrahydropyrrolo[3,4-c]pyrazol-3-yl)benzonitrile(100.0 mg, 0.32 mmol), 1-bromo-4-fluoro-5-isopropyl-2-methylbenzene(180.0 mg, 0.78 mmol), Pd(OAc)₂ (30.0 mg, 0.13 mmol), X-Phos (150.0 mg,0.33 mmol) and NaOtBu (100.0 mg, 1.0 mmol) in toluene (5 mL) was stirredat 110° C. for 1 h under N₂. The mixture was cooled to room temperature,diluted with water, and extracted with EtOAc. The organic layer wasseparated, washed with brine, dried over MgSO₄, and filtered. Thesolvent was concentrated under reduced pressure and the residue waspurified by silica gel flash chromatography (5 to 25% EtOAc in hexanes)to yield4-(2-(2,6-dimethylphenyl)-5-(4-fluoro-5-isopropyl-2-methylphenyl)-2,4,5,6-tetrahydropyrrolo[3,4-c]pyrazol-3-yl)benzonitrile.MS: (ES) m/z calculated for C₃₀H₃₀FN₄ [M+H]⁺ 465.2, found 465.2.

Step f: To a mixture of4-(2-(2,6-dimethylphenyl)-5-(4-fluoro-5-isopropyl-2-methylphenyl)-2,4,5,6-tetrahydropyrrolo[3,4-c]pyrazol-3-yl)benzonitrile(30.0 mg, 0.06 mmol) in DCM (1 mL) and DMSO (6 mL) was added 4 N aqueousNaOH (1.0 mL, 4.0 mmol) and H₂O₂ (0.40 mL, 35% in water). The mixturewas stirred for 30 min at room temperature, diluted with water, andextracted with EtOAc. The organic layer was separated, washed withbrine, dried over Na₂SO₄, and filtered. The solvent was concentratedunder reduced pressure and the residue was purified by preparative TLC(50% EtOAc in hexanes) to yield4-(2-(2,6-dimethylphenyl)-5-(4-fluoro-5-isopropyl-2-methylphenyl)-2,4,5,6-tetrahydropyrrolo[3,4-c]pyrazol-3-yl)benzamide.¹H NMR (400 MHz, CDCl₃): δ 7.63-7.71 (m, 2H), 7.20-7.31 (m, 1H),7.09-7.18 (m, 5H), 6.84-6.91 (m, 1H), 6.00 (br s, 1H), 5.58 (br s, 1H),4.58 (s, 2H), 4.49 (s, 2H), 3.14-3.26 (m, 1H), 2.39 (s, 3H), 2.02 (s,6H), 1.27 (d, J=7.0 Hz, 6H). MS: (ES) m/z calculated for C₃₀H₃₂FN₄O[M+H]⁺ 483.3, found 483.2.

Example 7

This example illustrates the evaluation of the biological activityassociated with specific compounds of the invention.

Materials and Methods

A. Cells

1. C5a Receptor Expressing Cells

a) U937 Cells

U937 cells are a monocytic cell line which express C5aR, and areavailable from ATCC (VA). These cells were cultured as a suspension inRPMI-1640 medium supplemented with 2 mM L-glutamine, 1.5 g/L sodiumbicarbonate, 4.5 g/L glucose, 10 mM HEPES, 1 mM sodium pyruvate, and 10%FBS. Cells were grown under 5% CO₂/95% air, 100% humidity at 37° C. andsubcultured twice weekly at 1:6 (cells were cultured at a density rangeof 1×10⁵ to 2×10⁶ cells/mL) and harvested at 1×10⁶ cells/mL. Prior toassay, cells are treated overnight with 0.5 mM of cyclic AMP (Sigma, OH)and washed once prior to use. cAMP treated U937 cells can be used inC5aR ligand binding and functional assays.

b) Isolated Human Neutrophils

Optionally, human or murine neutrophils can be used to assay forcompound activity. Neutrophils may be isolated from fresh human bloodusing density separation and centrifugation. Briefly, whole blood isincubated with equal parts 3% dextran and allowed to separate for 45minutes. After separation, the top layer is layered on top of 15 mls ofFicoll (15 mls of Ficoll for every 30 mls of blood suspension) andcentrifuged for 30 minutes at 400×g with no brake. The pellet at thebottom of the tube is then isolated and resuspended into PharmLyse RBCLysis Buffer (BD Biosciences, San Jose, Calif.) after which the sampleis again centrifuged for 10 minutes at 400×g with brake. The remainingcell pellet is resuspended as appropriate and consists of isolatedneutrophils.

B. Assays

1. Inhibition of C5aR Ligand Binding

cAMP treated U937 cells expressing C5aR were centrifuged and resuspendedin assay buffer (20 mM HEPES pH 7.1, 140 mM NaCl, 1 mM CaCl₂, 5 mMMgCl₂, and with 0.1% bovine serum albumin) to a concentration of 3×10⁶cells/mL. Binding assays were set up as follows. 0.1 mL of cells wasadded to the assay plates containing 5 μL of the compound, giving afinal concentration of ˜2-10 μM each compound for screening (or part ofa dose response for compound IC₅₀ determinations). Then 0.1 mL of ¹²⁵Ilabeled C5a (obtained from Perkin Elmer Life Sciences, Boston, Mass.)diluted in assay buffer to a final concentration of ˜50 pM, yielding˜30,000 cpm per well, was added, the plates sealed and incubated forapproximately 3 hours at 4° C. on a shaker platform. Reactions wereaspirated onto GF/B glass filters pre-soaked in 0.3% polyethyleneimine(PEI) solution, on a vacuum cell harvester (Packard Instruments;Meriden, Conn.). Scintillation fluid (40 μl; Microscint 20, PackardInstruments) was added to each well, the plates were sealed andradioactivity measured in a Topcount scintillation counter (PackardInstruments). Control wells containing either diluent only (for totalcounts) or excess C5a (1 μg/mL, for non-specific binding) were used tocalculate the percent of total inhibition for compound. The computerprogram Prism from GraphPad, Inc. (San Diego, Ca) was used to calculateIC₅₀ values. IC₅₀ values are those concentrations required to reduce thebinding of radiolabeled C5a to the receptor by 50%. (For furtherdescriptions of ligand binding and other functional assays, seeDairaghi, et al., J. Biol. Chem. 274:21569-21574 (1999), Penfold, etal., Proc. Natl. Acad. Sci. USA. 96:9839-9844 (1999), and Dairaghi, etal., J. Biol. Chem. 272:28206-28209 (1997)).

2. Calcium Mobilization

Optionally, compounds may be further assayed for their ability toinhibit calcium flux in cells. To detect the release of intracellularstores of calcium, cells (e.g., cAMP stimulated U937 or neutrophils) areincubated with 3 μM of INDO-1AM dye (Molecular Probes; Eugene, Oreg.) incell media for 45 minutes at room temperature and washed with phosphatebuffered saline (PBS). After INDO-1AM loading, the cells are resuspendedin flux buffer (Hank's balanced salt solution (HBSS) and 1% FBS).Calcium mobilization is measured using a Photon Technology Internationalspectrophotometer (Photon Technology International; New Jersey) withexcitation at 350 nm and dual simultaneous recording of fluorescenceemission at 400 nm and 490 nm. Relative intracellular calcium levels areexpressed as the 400 nm/490 nm emission ratio. Experiments are performedat 37° C. with constant mixing in cuvettes each containing 10⁶ cells in2 mL of flux buffer. The chemokine ligands may be used over a range from1 to 100 nM. The emission ratio is plotted over time (typically 2-3minutes). Candidate ligand blocking compounds (up to 10 μM) are added at10 seconds, followed by chemokines at 60 seconds (i.e., C5a; R&DSystems; Minneapolis, Minn.) and control chemokine (i.e., SDF-1α; R&DSystems; Minneapolis, Minn.) at 150 seconds.

3. Chemotaxis Assays

Optionally, compounds may be further assayed for their ability toinhibit chemotaxis in cells. Chemotaxis assays are performed using 5 μmpore polycarbonate, polyvinylpyrrolidone-coated filters in 96-wellchemotaxis chambers (Neuroprobe; Gaithersburg, Md.) using chemotaxisbuffer (Hank's balanced salt solution (HBSS) and 1% FBS). C5aR ligands(i.e., C5a, R&D Systems; Minneapolis, Minn.) are use to evaluatecompound mediated inhibition of C5aR mediated migration. Otherchemokines (i.e., SDF-1α; R&D Systems; Minneapolis, Minn.) are used asspecificity controls. The lower chamber is loaded with 29 μl ofchemokine (i.e., 0.03 nM C5a) and varying amounts of compound; the topchamber contains 100,000 U937 or neutrophil cells in 20 μl. The chambersare incubated 1.5 hours at 37° C., and the number of cells in the lowerchamber quantified either by direct cell counts in five high poweredfields per well or by the CyQuant assay (Molecular Probes), afluorescent dye method that measures nucleic acid content andmicroscopic observation.

C. Identification of Inhibitors of C5aR

1. Assay

To evaluate small organic molecules that prevent the C5a receptor frombinding ligand, an assay was employed that detected radioactive ligand(i.e, C5a) binding to cells expressing C5aR on the cell surface (forexample, cAMP stimulated U937 cells or isolated human neutrophils). Forcompounds that inhibited binding, whether competitive or not, fewerradioactive counts are observed when compared to uninhibited controls.

Equal numbers of cells were added to each well in the plate. The cellswere then incubated with radiolabeled C5a. Unbound ligand was removed bywashing the cells, and bound ligand was determined by quantifyingradioactive counts. Cells that were incubated without any organiccompound gave total counts; non-specific binding was determined byincubating the cells with unlabeled ligand and labeled ligand. Percentinhibition was determined by the equation:% inhibition=(1−[(sample cpm)−(nonspecific cpm)]/[(totalcpm)−(nonspecific cpm)])×100.

2. Dose Response Curves

To ascertain a candidate compound's affinity for C5aR as well as confirmits ability to inhibit ligand binding, inhibitory activity was titeredover a 1×10⁻¹⁰ to 1×10⁻⁴ M range of compound concentrations. In theassay, the amount of compound was varied; while cell number and ligandconcentration were held constant.

D. In Vivo Efficacy Models

The compounds of interest can be evaluated for potential efficacy intreating a C5a mediated conditions by determining the efficacy of thecompound in an animal model. In addition to the models described below,other suitable animal models for studying the compound of interest canbe found in Mizuno, M. et al., Expert Opin. Investig. Drugs (2005),14(7), 807-821, which is incorporated herein by reference in itsentirety.

1. Models of C5a Induced Leukopenia

a) C5a Induced Leukopenia in a Human C5aR Knock-in Mouse Model

To study the efficacy of compounds of the instant invention in an animalmodel, a recombinant mouse can be created using standard techniques,wherein the genetic sequence coding for the mouse C5aR is replaced withsequence coding for the human C5aR, to create a hC5aR-KI mouse. In thismouse, administration of hC5a leads to upregulation of adhesionmolecules on blood vessel walls which bind blood leukocytes,sequestering them from the blood stream. Animals are administered 20ug/kg of hC5a and 1 minute later leukocytes are quantified in peripheralblood by standard techniques. Pretreatment of mice with varying doses ofthe present compounds can almost completely block the hC5a inducedleukopenia.

b) C5a Induced Leukopenia in a Cynomolgus Model

To study the efficacy of compounds of the instant invention in anon-human primate model model, C5a induced leucopenia is studied in acynomolgus model. In this model administration of hC5a leads toupregulation of adhesion molecules on blood vessel walls which bindblood leukocytes, hence sequestering them from the blood stream. Animalsare administered 10 ug/kg of hC5a and 1 minute later leukocytes arequantified in peripheral blood.

Mouse Model of ANCA Induced Vasculitis

On day 0 hC5aR-KI mice are intraveneously injected with 50 mg/kgpurified antibody to myeloperoxidase (Xiao et al, J. Clin. Invest. 110:955-963 (2002)). Mice are further dosed with oral daily doses ofcompounds of the invention or vehicle for seven days, then mice aresacrificed and kidneys collected for histological examination. Analysisof kidney sections can show significantly reduced number and severity ofcrescentic and necrotic lesions in the glomeruli when compared tovehicle treated animals.

2. Mouse Model of Choroidal Neovascularization

To study the efficacy of compounds of the instant invention in treatmentof age related macular degeneration (AMD) the bruch membrane in the eyesof hC5aR-KI mice are ruptured by laser photocoagulation (Nozika et al,PNAS 103: 2328-2333 (2006). Mice are treated with vehicle or a dailyoral or appropriate intra-vitreal dose of a compound of the inventionfor one to two weeks. Repair of laser induced damage andneovascularization are assessed by histology and angiography.

3. Rheumatoid Arthritis Models

a) Rabbit Model of Destructive Joint Inflammation

To study the effects of candidate compounds on inhibiting theinflammatory response of rabbits to an intra-articular injection of thebacterial membrane component lipopolysaccharide (LPS), a rabbit model ofdestructive joint inflammation is used. This study design mimics thedestructive joint inflammation seen in arthritis. Intra-articularinjection of LPS causes an acute inflammatory response characterized bythe release of cytokines and chemokines, many of which have beenidentified in rheumatoid arthritic joints. Marked increases inleukocytes occur in synovial fluid and in synovium in response toelevation of these chemotactic mediators. Selective antagonists ofchemokine receptors have shown efficacy in this model (see Podolin, etal., J. Immunol. 169(11):6435-6444 (2002)).

A rabbit LPS study is conducted essentially as described in Podolin, etal. ibid., female New Zealand rabbits (approximately 2 kilograms) aretreated intra-articularly in one knee with LPS (10 ng) together witheither vehicle only (phosphate buffered saline with 1% DMSO) or withaddition of candidate compound (dose 1=50 μM or dose 2=100 μM) in atotal volume of 1.0 mL. Sixteen hours after the LPS injection, knees arelavaged and cells counts are performed. Beneficial effects of treatmentwere determined by histopathologic evaluation of synovial inflammation.Inflammation scores are used for the histopathologic evaluation:1—minimal, 2—mild, 3—moderate, 4—moderate-marked.

b) Evaluation of a Compound in a Rat Model of Collagen Induced Arthritis

A 17 day developing type II collagen arthritis study is conducted toevaluate the effects of a candidate compound on arthritis inducedclinical ankle swelling. Rat collagen arthritis is an experimental modelof polyarthritis that has been widely used for preclinical testing ofnumerous anti-arthritic agents (see Trentham, et al., J. Exp. Med.146(3):857-868 (1977), Bendele, et al., Toxicologic Pathol. 27:134-142(1999), Bendele, et al., Arthritis Rheum. 42:498-506 (1999)). Thehallmarks of this model are reliable onset and progression of robust,easily measurable polyarticular inflammation, marked cartilagedestruction in association with pannus formation and mild to moderatebone resorption and periosteal bone proliferation.

Female Lewis rats (approximately 0.2 kilograms) are anesthetized withisoflurane and injected with Freund's Incomplete Adjuvant containing 2mg/mL bovine type II collagen at the base of the tail and two sites onthe back on days 0 and 6 of this 17 day study. A candidate compound isdosed daily in a sub-cutaneous manner from day 0 till day 17 at aefficacious dose. Caliper measurements of the ankle joint diameter weretaken, and reducing joint swelling is taken as a measure of efficacy.

4. Rat Model of Sepsis

To study the effect of compounds of interest on inhibiting thegeneralized inflammatory response that is associated with a sepsis likedisease, the Cecal Ligation and Puncture (CLP) rat model of sepsis isused. A Rat CLP study is conducted essentially as described in FujimuraN, et al. (American Journal Respiratory Critical Care Medicine 2000;161: 440-446). Briefly described here, Wistar Albino Rats of both sexesweighing between 200-250 g are fasted for twelve hours prior toexperiments. Animals are kept on normal 12 hour light and dark cyclesand fed standard rat chow up until 12 hours prior to experiment. Thenanimals are split into four groups; (i) two sham operation groups and(ii) two CLP groups. Each of these two groups (i.e., (i) and (ii)) issplit into vehicle control group and test compound group. Sepsis isinduced by the CLP method. Under brief anesthesia a midline laparotomyis made using minimal dissection and the cecum is ligated just below theileocaecal valve with 3-0 silk, so the intestinal continuity ismaintained. The antimesinteric surface of the cecum is perforated withan 18 gauge needle at two locations 1 cm apart and the cecum is gentlysqueezed until fecal matter is extruded. The bowel is then returned tothe abdomen and the incision is closed. At the end of the operation, allrats are resuscitated with saline, 3 ml/100 g body weight, givensubcutaneously. Postoperatively, the rats are deprived of food, but havefree access to water for the next 16 hours until they are sacrificed.The sham operated groups are given a laparotomy and the cecum ismanipulated but not ligated or perforated. Beneficial effects oftreatment are measured by histopathological scoring of tissues andorgans as well as measurement of several key indicators of hepaticfunction, renal function, and lipid peroxidation. To test for hepaticfunction aspartate transaminase (AST) and alanine transaminase (ALT) aremeasured. Blood urea nitrogen and creatinine concentrations are studiedto assess renal function. Pro-inflammatory cytokines such as TNF-alphaand IL-1beta are also assayed by ELISA for serum levels.

5. Mouse SLE Model of Experimental Lupus Nephritis.

To study the effect of compounds of interest on a Systemic LupusErythematosus (SLE), the MRL/lpr murine SLE model is used. TheMRL/Mp-Tmfrsf6^(lpr/lpr) strain (MRL/lpr) is a commonly used mouse modelof human SLE. To test compounds efficacy in this model male MRL/lpr miceare equally divided between control and C5aR antagonists groups at 13weeks of age. Then over the next 6 weeks compound or vehicle isadministered to the animals via osmotic pumps to maintain coverage andminimize stress effects on the animals. Serum and urine samples arecollected bi-weekly during the six weeks of disease onset andprogression. In a minority of these mice glomerulosclerosis developsleading to the death of the animal from renal failure. Followingmortality as an indicator of renal failure is one of the measuredcriteria and successful treatment will usually result in a delay in theonset of sudden death among the test groups. In addition, the presenceand magnitude of renal disease may also be monitored continuously withblood urea nitrogen (BUN) and albuminuria measurements. Tissues andorgans were also harvested at 19 weeks and subjected to histopathologyand immunohistochemistry and scored based on tissue damage and cellularinfiltration.

6. Rat Model of COPD

Smoke induced airway inflammation in rodent models may be used to assessefficacy of compounds in Chronic Obstructive Pulmonary Disease (COPD).Selective antagonists of chemokines have shown efficacy in this model(see, Stevenson, et al., Am. J. Physiol Lung Cell Mol Physiol. 288L514-L522, (2005)). An acute rat model of COPD is conducted as describedby Stevenson et al. A compound of interest is administered eithersystemically via oral or IV dosing; or locally with nebulized compound.Male Sprague-Dawley rats (350-400 g) are placed in Perspex chambers andexposed to cigarette smoke drawn in via a pump (50 mL every 30 secondswith fresh air in between). Rats are exposed for a total period of 32minutes. Rats are sacrificed up to 7 days after initial exposure. Anybeneficial effects of treatment are assessed by a decrease inflammatorycell infiltrate, decreases in chemokine and cytokine levels.

In a chronic model, mice or rats are exposed to daily tobacco smokeexposures for up to 12 months. Compound is administered systemically viaonce daily oral dosing, or potentially locally via nebulized compound.In addition to the inflammation observed with the acute model (Stevensenet al.), animals may also exhibit other pathologies similar to that seenin human COPD such as emphysema (as indicated by increased mean linearintercept) as well as altered lung chemistry (see Martorana et al, Am.J. Respir. Crit Care Med. 172(7): 848-53.

7. Mouse EAE Model of Multiple Sclerosis

Experimental autoimmune encephalomyelitis (EAE) is a model of humanmultiple sclerosis. Variations of the model have been published, and arewell known in the field. In a typical protocol, C57BL/6 (Charles RiverLaboratories) mice are used for the EAE model. Mice are immunized with200 ug myelin oligodendrocyte glycoprotein (MOG) 35-55 (PeptideInternational) emulsified in Complete Freund's Adjuvant (CFA) containing4 mg/ml Mycobacterium tuberculosis (Sigma-Aldrich) s.c. on day 0. Inaddition, on day 0 and day 2 animals are given 200 ng of pertussis toxin(Calbiochem) i.v. Clinical scoring is based on a scale of 0-5: 0, nosigns of disease; 1, flaccid tail; 2, hind limb weakness; 3, hind limbparalysis; 4, forelimb weakness or paralysis; 5, moribund. Dosing of thecompounds of interest to be assessed can be initiated on day 0(prophylactic) or day 7 (therapeutic, when histological evidence ofdisease is present but few animals are presenting clinical signs) anddosed once or more per day at concentrations appropriate for theiractivity and pharmacokinetic properties, e.g. 100 mg/kg s.c. Efficacy ofcompounds can be assessed by comparisons of severity (maximum meanclinical score in presence of compound compared to vehicle), or bymeasuring a decrease in the number of macrophages (F4/80 positive)isolated from spinal cords. Spinal cord mononuclear cells can beisolated via discontinuous Percoll-gradient. Cells can be stained usingrat anti-mouse F4/80-PE or rat IgG2b-PE (Caltag Laboratories) andquantitated by FACS analysis using 10 ul of Polybeads per sample(Polysciences).

8. Mouse Model of Kidney Transplantation

Transplantation models can be performed in mice, for instance a model ofallogenic kidney transplant from C57BL/6 to BALB/c mice is described inFaikah Gueler et al, JASN Express, Aug. 27, 2008. Briefly, mice areanesthetized and the left donor kidney attached to a cuff of the aortaand the renal vein with a small caval cuff, and the ureters removed enblock. After left nephrectomy of the recipient, the vascular cuffs areanastomosed to the recipient abdominal aorta and vena cava,respectively, below the level of the native renal vessels. The ureter isdirectly anastomosed into the bladder. Cold ischemia time is 60 min, andwarm ischemia time is 30 min. The right native kidney can be removed atthe time of allograft transplantation or at posttransplantation day 4for long-term survival studies. General physical condition of the miceis monitored for evidence of rejection. Compound treatment of animalscan be started before surgery or immediately after transplantation, egby sub cut injection once daily. Mice are studied for renal function andsurvival. Serum creatinine levels are measured by an automated method(Beckman Analyzer, Krefeld, Germany).

9. Mouse Model of Ischemia/Reperfusion

A mouse model of ischemia/reperfusion injury can be performed asdescribed by Xiufen Zheng et al, Am. J. Pathol, Vol 173:4, October,2008. Briefly, CD1 mice aged 6-8 weeks are anesthetized and placed on aheating pad to maintain warmth during surgery. Following abdominalincisions, renal pedicles are bluntly dissected and a microvascularclamp placed on the left renal pedicle for 25-30 minutes. Followingischemia the clamps are removed along with the right kidney, incisionssutured, and the animals allowed to recover. Blood is collected forserum creatinine and BUN analysis as an indicator of kidney health.Alternatively animal survival is monitored over time. Compound can beadministered to animals before and/or after the surgery and the effectson serum creatinine, BUN or animal survival used as indicators ofcompound efficacy.

10. Mouse Model of Tumor Growth

C57BL/6 mice 6-16 weeks of age are injected subcutaneously with 1×105TC-1 cells (ATCC, VA) in the right or left rear flank. Beginning about 2weeks after cell injection, tumors are measured with calipers every 2-4d until the tumor size required the mice are killed. At the time ofsacrifice animals are subjected to a full necropsy and spleens andtumors removed. Excised tumors are measured and weighed. Compounds maybe administered before and/or after tumor injections, and a delay orinhibition of tumor growth used to assess compound efficacy.

Example 8

The compounds in Table 1, below, were prepared using the methodsdescribed above. Characterization data is provided for each compoundlisted. Activity is provided as follows for the chemotaxis assay usingU937 cells as described herein (Example 7): +, 500 nM<IC₅₀; ++, 50nM<IC₅₀≤500 nM; +++, 5 nM<IC₅₀≤50 nM; and ++++, IC₅₀≤5 nM.

TABLE 1 Structure, Characterization Data and Biological Activity Data ofSpecific Embodiments Compound MS: ES (m/z) Mig IC₅₀ Number Structure[M + H]⁺ (nM) 1.001

682.2 ++++ 1.002

678.3 ++++ 1.003

662.5 ++++ 1.004

564.3 ++++ 1.005

633.3 + 1.006

570.6 ++++ 1.007

638.6 ++++ 1.008

520.3 +++ 1.009

594.6 ++++ 1.010

662.5 ++++ 1.011

598.6 ++++ 1.012

524.5 ++++ 1.013

524.5 ++++ 1.014

598.3 ++++ 1.015

538.6 ++++ 1.016

510.6 +++ 1.017

666.2 ++++ 1.018

666.2 ++++ 1.019

446.5 ++ 1.020

525.2 + 1.021

508.4 + 1.022

418.4 ++ 1.023

507.3 ++ 1.024

507.3 +++ 1.025

507.3 + 1.026

403.2 + 1.027

466.5 + 1.028

466.5 + 1.029

448.3 + 1.030

493.3 +++ 1.031

493.3 +++ 1.032

463.2 + 1.033

432.2 + 1.034

469.2 ++ 1.035

483.2 +++ 1.036

493.3 + 1.037

437.2 + 1.038

433.4 + 1.039

491.4 + 1.040

493.3 ++ 1.041

537.5 ++++ 1.042

547.5 + 1.043

480.3 + 1.044

482.3 ++ 1.045

466.3 ++ 1.046

423.3 ++ 1.047

479.3 ++++ 1.048

494.5 ++ 1.049

494.3 ++ 1.050

465.3 + 1.051

451.3 ++++ 1.052

510.5 + 1.053

495.5 +++ 1.054

481.3 +++ 1.055

467.3 +++

While particular embodiments of this invention are described herein,upon reading the description, variations of the disclosed embodimentsmay become apparent to individuals working in the art, and it isexpected that those skilled artisans may employ such variations asappropriate. Accordingly, it is intended that the invention be practicedotherwise than as specifically described herein, and that the inventionincludes all modifications and equivalents of the subject matter recitedin the claims appended hereto as permitted by applicable law. Moreover,any combination of the above-described elements in all possiblevariations thereof is encompassed by the invention unless otherwiseindicated herein or otherwise clearly contradicted by context.

All publications, patent applications, accession numbers, and otherreferences cited in this specification are herein incorporated byreference as if each individual publication or patent application werespecifically and individually indicated to be incorporated by reference.

What is claimed is:
 1. A compound of Formula (I)

or a pharmaceutically acceptable salt thereof, wherein, ring vertex a isN or C(R^(2c)), ring vertex b is N or C(R^(2d)), and ring vertex e is Nor C(R^(2e)), wherein no more than one of a, b and e is N; X¹ isselected from the group consisting of a bond, C₁₋₈ alkylene, C(O),C(O)—C₁₋₄ alkylene, and S(O)₂; R¹ is selected from the group consistingof a) 5- to 10-membered heteroaryl having from 1 to 4 heteroatoms asring vertices selected from N, O and S; b) C₆₋₁₀ aryl; c) C₃₋₈cycloalkyl; d) 4- to 8-membered heterocycloalkyl having from 1 to 2heteroatoms as ring vertices selected from N, O and S; and e) C₁₋₈alkyl, C₁₋₈ alkoxy, C₁₋₈ haloalkyl, —C(O)NR^(1a)R^(1b), and —CO₂R^(1a);wherein R^(1a) and R^(1b) are each independently selected from the groupconsisting of hydrogen, C₁₋₈ alkyl, C₆₋₁₀ aryl, and —C₁₋₆ alkylene-C₆₋₁₀aryl; wherein the group —X¹—R¹ is optionally substituted with 1 to 5R^(x) substituents; R^(2a) and R^(2e) are each independently selectedfrom the group consisting of hydrogen, C₁₋₆ alkyl, C₁₋₆ alkoxy, C₁₋₆haloalkyl, —O—C₁₋₆ haloalkyl, —S—C₁₋₆ alkyl, —C₁₋₆ alkyl-O—C₁₋₆ alkyl,—C₁₋₆ alkyl-S—C₁₋₆ alkyl, CN, and halogen, and at least one of R^(2a)and R^(2e) is other than hydrogen; R^(2b), R^(2c), and R^(2d) are eachindependently selected from the group consisting of hydrogen, C₁₋₆alkyl, C₁₋₆ alkoxy, C₁₋₆ haloalkyl, —O—C₁₋₆ haloalkyl, —S—C₁₋₆ alkyl,—C₁₋₆ alkyl-O—C₁₋₆ alkyl, —C₁₋₆ alkyl-S—C₁₋₆ alkyl, cyano, and halogen;each R³ is independently selected from the group consisting of hydroxyl,C₁₋₄ alkyl, C₁₋₄ haloalkyl and C₁₋₄ hydroxyalkyl, and optionally two R³groups on the same carbon atom are combined to form oxo (═O), andoptionally two R³ groups and the carbon atoms they are attached to forma 3-6 membered ring with 0-2 hetereoatoms as ring members selected fromO, N, and S; R⁴ is independently selected from the group consisting of

each R⁵ is independently selected from the group consisting of C₁₋₈alkyl, C₁₋₈ alkoxy, C₁₋₈ haloalkyl, C₁₋₈ haloalkoxy, C₁₋₈ hydroxyalkyl,halogen, OH, CN, C(O)R^(5a) and CO₂R^(5a); wherein each R^(5a) isindependently selected from the group consisting of hydrogen, C₁₋₄alkyl, and C₁₋₄ haloalkyl; each R^(x) is independently selected from thegroup consisting of halogen, CN, C₁₋₄ alkyl, C₁₋₄ alkoxy, C₁₋₄haloalkyl, C₁₋₄ haloalkoxy, C₁₋₄ hydroxy, C₂₋₄ alkenyl, C₃₋₆ cycloalkyl,CO₂—C₁₋₄ alkyl, and CONH₂; the subscript m is 0, 1, 2, 3 or 4; and thesubscript n is 0, 1, 2 or
 3. 2. The compound of claim 1, or apharmaceutically acceptable salt thereof, wherein R⁴ is selected fromthe group consisting of


3. The compound of claim 1, or a pharmaceutically acceptable saltthereof, wherein R⁴ is selected from the group consisting of


4. The compound of claim 1, or a pharmaceutically acceptable saltthereof, wherein X¹ is a bond.
 5. The compound of claim 1, or apharmaceutically acceptable salt thereof, wherein X¹ is C(O).
 6. Thecompound of claim 1, or a pharmaceutically acceptable salt thereof,wherein X¹ is C₁₋₈ alkylene.
 7. The compound of claim 1, or apharmaceutically acceptable salt thereof, wherein X¹ is C(O)—C₁₋₄alkylene or S(O)₂.
 8. The compound of claim 1, or a pharmaceuticallyacceptable salt thereof, wherein R¹ is a 5- to 10-membered heteroarylhaving from 1 to 4 heteroatoms as ring vertices selected from N, O andS; and wherein the group —X¹—R¹ is optionally substituted with 1 to 4R^(x) substituents.
 9. The compound of claim 8, wherein R¹ is selectedfrom the group consisting of pyrazolyl, pyridyl, pyrimidinyl,imidazolyl, thiazolyl, thiadiazolyl and pyrazinyl; and wherein the group—X¹—R¹ is optionally substituted with 1 to 4 R^(x) substituents.
 10. Thecompound of claim 1, or a pharmaceutically acceptable salt thereof,having formula (Ia) or (Ib):


11. The compound of claim 1, or a pharmaceutically acceptable saltthereof, wherein R¹ is C₆₋₁₀ aryl; and wherein the group —X¹—R¹ isoptionally substituted with 1 to 4 R^(x) substituents.
 12. The compoundof claim 11, wherein R¹ is phenyl; and wherein the group —X¹—R¹ isoptionally substituted with 1 to 4 R^(x) substituents.
 13. The compoundof claim 1, or a pharmaceutically acceptable salt thereof, wherein R¹ isC₃₋₈ cycloalkyl; and wherein the group —X¹—R¹ is optionally substitutedwith 1 to 4 R^(x) substituents.
 14. The compound of claim 13, wherein R¹is selected from the group consisting of cyclobutyl, cyclopentyl andcyclohexyl; and wherein the group —X¹—R¹ is optionally substituted with1 to 4 R^(x) substituents.
 15. The compound of claim 1, or apharmaceutically acceptable salt thereof, wherein R¹ is a 4- to8-membered heterocycloalkyl having from 1 to 2 heteroatoms as ringvertices selected from N, O and S; and wherein the group —X¹—R¹ isoptionally substituted with 1 to 4 R^(x) substituents.
 16. The compoundof claim 15, wherein R¹ is selected from the group consisting ofoxetanyl, tetrahydrofuranyl, tetrahydropyranyl and morpholinyl; andwherein the group —X¹—R¹ is optionally substituted with 1 to 4 R^(x)substituents.
 17. The compound of claim 1, or a pharmaceuticallyacceptable salt thereof, wherein R¹ is selected from the groupconsisting of C₁₋₈ alkyl, C₁₋₈ alkoxy, C₁₋₈ haloalkyl,—C(O)NR^(1a)R^(1b), and —CO₂R^(1a); wherein R^(1a) and R^(1b) are eachindependently selected from the group consisting of hydrogen, C₁₋₈alkyl, C₆₋₁₀ aryl, and —C₁₋₆ alkylene-C₆₋₁₀ aryl; and wherein the group—X¹—R¹ is optionally substituted with 1 to 4 R^(x) substituents.
 18. Thecompound of claim 1, or a pharmaceutically acceptable salt thereof,wherein R¹ is selected from the group consisting of phenyl, pyridyl,pyrimidinyl, and pyrazinyl; and wherein the group —X¹—R¹ is optionallysubstituted with 1 to 4 R^(x) substituents.
 19. The compound of claim 1,or a pharmaceutically acceptable salt thereof, wherein ring vertices aand b are CH; R^(2b) is H; ring vertex e is C(R^(2e)), and R^(2a) andR^(2e) are independently selected from the group consisting of C₁₋₆alkyl, C₁₋₆ alkoxy, C₁₋₆ haloalkyl, —O—C₁₋₆ haloalkyl, —S—C₁₋₆ alkyl,—C₁₋₆ alkyl-O—C₁₋₆ alkyl, —C₁₋₆ alkyl-S—C₁₋₆ alkyl, CN, and halogen. 20.The compound of claim 1, or a pharmaceutically acceptable salt thereof,wherein ring vertices a and b are CH; R^(2b) is H; ring vertex e isC(R^(2e)), and R^(2a) and R^(2e) are independently selected from thegroup consisting of C₁₋₆ alkyl, C₁₋₆ alkoxy and halogen.
 21. Thecompound of claim 1, or a pharmaceutically acceptable salt thereof,wherein n is 0, 1 or 2 and each R⁵, when present, is independentlyselected from the group consisting of F, Cl, CN, C₁₋₄ alkyl and C₁₋₄alkoxy.
 22. The compound of claim 1, or a pharmaceutically acceptablesalt thereof, wherein n is 0, 1 or 2 and each R⁵, when present, isindependently selected from the group consisting of F, Cl, CN, CH₃ andOCH₃.
 23. The compound of claim 1, or a pharmaceutically acceptable saltthereof, wherein m is 0, 1 or 2 and each R³, when present, is C₁₋₄alkyl.
 24. The compound of claim 1, or a pharmaceutically acceptablesalt thereof, wherein R¹ is selected from the group consisting of phenylor pyridyl, wherein the group —X¹—R¹ is optionally substituted with 1 to4 R^(x) substituents; ring vertices a and b are CH; R^(2b) is H; ringvertex e is C(R^(2e)), and R^(2a) and R^(2e) are independently selectedfrom the group consisting of C₁₋₆ alkyl, C₁₋₆ alkoxy and halogen; m is0, 1 or 2 and each R³, when present, is CH₃; R⁴ is selected from thegroup consisting of

n is 0, 1 or 2 and each R⁵, when present, is selected from the groupconsisting of F, Cl, CN, CH₃ and OCH₃.
 25. The compound of claim 1, or apharmaceutically acceptable salt thereof, wherein —X¹—R¹ is selectedfrom the group consisting of:


26. The compound of claim 1, or a pharmaceutically acceptable saltthereof, wherein

is selected from the group consisting of


27. The compound of claim 1, or a pharmaceutically acceptable saltthereof, wherein n is
 0. 28. The compound of claim 1, or apharmaceutically acceptable salt thereof, wherein m is 2 and the two R³groups are on the same carbon atom and are combined to form oxo (═O).29. The compound of claim 1, or a pharmaceutically acceptable saltthereof, wherein said compound is selected from the group consisting of:


30. A pharmaceutical composition comprising a compound of claim 1 or apharmaceutically acceptable salt thereof and a pharmaceuticallyacceptable carrier.
 31. The pharmaceutical composition of claim 30,formulated for oral, intravenous, transdermal or subcutaneousadministration.
 32. The pharmaceutical composition of claim 30, furthercomprising one or more additional therapeutic agents.
 33. Thepharmaceutical composition of claim 32, wherein the one or moreadditional therapeutic agent is selected from the group consisting ofcorticosteroids, steroids, immunosuppressants, Immunoglobulin Gagonists, Dipeptidyl peptidase IV inhibitors, Lymphocyte functionantigen-3 receptor antagonists, Interleukin-2 ligands, Interleukin-1beta ligand inhibitors, IL-2 receptor alpha subunit inhibitors, HGF genestimulators, IL-6 antagonists, IL-5 antagonists, Alpha 1 antitrypsinstimulators, Cannabinoid receptor antagonists, Histone deacetylaseinhibitors, AKT protein kinase inhibitors, CD20 inhibitors, Abl tyrosinekinase inhibitors, JAK tyrosine kinase inhibitors, TNF alpha ligandinhibitors, Hemoglobin modulators, TNF antagonists, proteasomeinhibitors, CD3 modulators, Hsp 70 family inhibitors, Immunoglobulinagonists, CD30 antagonists, tubulin antagonists, Sphingosine-1-phosphatereceptor-1 agonists, connective tissue growth factor ligand inhibitors,caspase inhibitors, adrenocorticotrophic hormone ligands, Btk tyrosinekinase inhibitors, Complement C1s subcomponent inhibitors,Erythropoietin receptor agonists, B-lymphocyte stimulator ligandinhibitors, Cyclin-dependent kinase-2 inhibitors, P-selectinglycoprotein ligand-1 stimulators, mTOR inhibitors, Elongation factor 2inhibitors, Cell adhesion molecule inhibitors, Factor XIII agonists,Calcineurin inhibitors, Immunoglobulin G1 agonists, Inosinemonophosphate dehydrogenase inhibitors, Complement C1s subcomponentinhibitors, Thymidine kinase modulators, Cytotoxic T-lymphocyteprotein-4 modulators, Angiotensin II receptor antagonists, AngiotensinII receptor modulators, TNF superfamily receptor 12A antagonists, CD52antagonists, Adenosine deaminase inhibitors, T-cell differentiationantigen CD6 inhibitors, FGF-7 ligands, dihydroorotate dehydrogenaseinhibitors, Syk tyrosine kinase inhibitors, Interferon type I receptorantagonists, Interferon alpha ligand inhibitors, Macrophage migrationinhibitory factor inhibitors, Integrin alpha-V/beta-6 antagonists,Cysteine protease stimulators, p38 MAP kinase inhibitors, TP53 geneinhibitors, Shiga like toxin I inhibitors, Fucosyltransferase 6stimulators, Interleukin 22 ligands, IRS1 gene inhibitors, Proteinkinase C stimulators, Protein kinase C alpha inhibitors, CD74antagonists, Immunoglobulin gamma Fc receptor IIB antagonists, T-cellantigen CD7 inhibitors, CD95 antagonists, N acetylmannosamine kinasestimulators, Cardiotrophin-1 ligands, Leukocyte elastase inhibitors,CD40 ligand receptor antagonists, CD40 ligand modulators, IL-17antagonists, TLR-2 antagonists, Mannan-binding lectin serine protease-2(MASP-2) inhibitors, Factor B inhibitors, Factor D inhibitors, C3aRmodulators, C5aR2 modulators, T cell receptor antagonists, PD-1inhibitors, PD-L1 inhibitors, TIGIT inhibitors, TIM-3 inhibitors, LAG-3inhibitors, VISTA inhibitors, STING agonists, IDO inhibitors, adenosinereceptor modulators, CD39 inhibitors, CD73 inhibitors, antagonists ofthe chemokine receptors, especially CXCR1, CXCR2, CXCR3, CXCR4, CXCR7,CCR1, CCR2, CCR3, CCR4, CCR5, CCR7, CCR7, CCR9, CX3CR1 and CXCR6, andcombinations thereof.
 34. The compound of claim 1, selected from thegroup consisting of

or a pharmaceutically acceptable salt thereof.
 35. The compound of claim1, having the structure:

or a pharmaceutically acceptable salt thereof.
 36. The compound of claim1, having the structure:

or a pharmaceutically acceptable salt thereof.
 37. The compound of claim1, having the structure:

or a pharmaceutically acceptable salt thereof.
 38. The compound of claim1, having the structure:

or a pharmaceutically acceptable salt thereof.